This page has only limited features, please log in for full access.

Dr. Avi Priel
Pharmacology section, The Institute for Drug Research (IDR), The School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, Israel

Basic Info


Research Keywords & Expertise

0 plant toxins
0 Venom toxins
0 Disulfide-rich peptides
0 Ion channels biophysics and pharmacology
0 Snake and spider toxins

Fingerprints

Venom toxins
Snake and spider toxins

Honors and Awards

The user has no records in this section


Career Timeline

The user has no records in this section.


Short Biography

The user biography is not available.
Following
Followers
Co Authors
The list of users this user is following is empty.
Following: 0 users

Feed

Brief research report article
Published: 12 June 2020 in Frontiers in Pharmacology
Reads 0
Downloads 0

Neuronal signals are processed along the nociceptive pathway to convey discriminative information, which would manifest in the produced pain sensation. The transient receptor potential vanilloid 1 (TRPV1), an important signaling complex in nociceptors termini, is activated by different noxious stimuli that underlie distinct pain sensations. For example, while endovanilloids are associated with inflammatory pain and hypersensitivity through TRPV1 activation, the exovanilloid toxin, capsaicin, evokes an acute pain by activating this channel. Differences in the TRPV1 activation profile evoked by exogenous and endogenous vanilloids were suggested to underlie this disparity in pain sensations. However, the cellular processes that lead to these differences in pain sensation mediated by the same channel are not fully understood. Here, we sought to describe the neuronal response of TRPV1-expressing nociceptors to exo-and endovanilloids. To this end, we performed current-clamp recordings in rat trigeminal neurons exposed to either capsaicin or intracellular endovanilloids produced downstream of the bradykinin receptor BK2. Our results show that lipoxygenase metabolites generate persistent TRPV1-dependent action potential firing while capsaicin evokes robust depolarization and high-frequency firing that is quickly terminated by depolarization block. Additionally, we found that a weak TRPV1 activation prolongs action potential firing. Overall, our results indicate different firing patterns evoked by inflammatory mediators and capsaicin via TRPV1 that correlate with the respective subsequent pain sensation. These findings also suggest that differences in neuronal activation stem from the variable degree of TRPV1 activation they produce.

ACS Style

Rakesh Kumar; Matan Geron; Adina Hazan; Avi Priel. Endogenous and Exogenous Vanilloids Evoke Disparate TRPV1 Activation to Produce Distinct Neuronal Responses. Frontiers in Pharmacology 2020, 11, 1 .

AMA Style

Rakesh Kumar, Matan Geron, Adina Hazan, Avi Priel. Endogenous and Exogenous Vanilloids Evoke Disparate TRPV1 Activation to Produce Distinct Neuronal Responses. Frontiers in Pharmacology. 2020; 11 ():1.

Chicago/Turabian Style

Rakesh Kumar; Matan Geron; Adina Hazan; Avi Priel. 2020. "Endogenous and Exogenous Vanilloids Evoke Disparate TRPV1 Activation to Produce Distinct Neuronal Responses." Frontiers in Pharmacology 11, no. : 1.

Original research article
Published: 17 January 2020 in Frontiers in Pharmacology
Reads 0
Downloads 0

C. elegans PVD neurons are conserved for morphology, function and molecular determinants with mammalian polymodal nociceptors. Functions of polymodal nociceptors require activities of multiple ion channels and receptors including members of the TRP family. GTL-1, a member of the TRPM subclass of TRP channels, was previously shown to amplify PVD-mediated responses to optogenetic stimuli. Here we characterize effects of GTL-1 on PVD-mediated behavioral responses to noxious stimuli. We show that GTL-1 is required within PVD for the immediate and enduring response to thermal (cold) stimuli. But, find no significant reduction in percent animals responding to single or to repeated noxious mechanical stimuli. Nevertheless, PVD specific knockdown of gtl-1expression reduces the magnitude of responses to noxious mechanical stimuli. To understand GTL-1’s mechanism of action we expressed it in HEK293 cells. Our results show GTL-1-dependent currents induced by activation of a Gαq-coupled Designer Receptor Exclusively Activated by Designer Drugs (DREADD). In addition, using excised patches we show that GTL-1 can be activated by internal calcium. Our results are consistent with indirect, calcium dependent, activation of GTL-1 by noxious stimuli. This mechanism explains the GTL-1-dependent amplification of responses to multiple stimuli optogenetic and sensory in PVD.

ACS Style

Emiliano Cohen; Rakesh Kumar; Tal Zinger; Avi Priel; Millet Treinin. GTL-1, a Calcium Activated TRPM Channel, Enhances Nociception. Frontiers in Pharmacology 2020, 10, 1567 .

AMA Style

Emiliano Cohen, Rakesh Kumar, Tal Zinger, Avi Priel, Millet Treinin. GTL-1, a Calcium Activated TRPM Channel, Enhances Nociception. Frontiers in Pharmacology. 2020; 10 ():1567.

Chicago/Turabian Style

Emiliano Cohen; Rakesh Kumar; Tal Zinger; Avi Priel; Millet Treinin. 2020. "GTL-1, a Calcium Activated TRPM Channel, Enhances Nociception." Frontiers in Pharmacology 10, no. : 1567.

Protocol
Published: 02 October 2019 in Methods in Molecular Biology
Reads 0
Downloads 0

Snake and spider envenomation have a considerable impact on public health. Their pathology is induced by a variety of toxins composing the venom which induce cytotoxicity to cells of different organs by several cell death pathways. Described in this chapter are methods in vitro used to assess venoms and toxin-induced cell death using mammalian cell cultures. The chapter is divided into five sections: (1) a brief overview of in vitro cytotoxicity and categories of cell death induced by venoms and toxins; (2) a common method to measure necrotic cell death using lactate dehydrogenase (LDH) release; (3) a flow cytometry method that simultaneously measures necrosis and apoptosis; (4) measurements of nuclear morphology; and (5) measurements of the autophagy following microtubule-associated protein light chain 3 (LC3) expression, by immunoblotting and by fluorescence microscopy of LC3-positive vesicles, to assess the levels of autophagosomes.

ACS Style

Yossi Maatuf; Avi Priel; Philip Lazarovici. Measurements of Cell Death Induced by Snake and Spider's Venoms and Derived Toxins. Methods in Molecular Biology 2019, 2068, 239 -268.

AMA Style

Yossi Maatuf, Avi Priel, Philip Lazarovici. Measurements of Cell Death Induced by Snake and Spider's Venoms and Derived Toxins. Methods in Molecular Biology. 2019; 2068 ():239-268.

Chicago/Turabian Style

Yossi Maatuf; Avi Priel; Philip Lazarovici. 2019. "Measurements of Cell Death Induced by Snake and Spider's Venoms and Derived Toxins." Methods in Molecular Biology 2068, no. : 239-268.

Protocol
Published: 02 October 2019 in Methods in Molecular Biology
Reads 0
Downloads 0

Many toxins from a variety of venomous animals and plants have evolved to target neuronal ion channels and receptors. However, a significant obstacle in the study of these toxins is the finding and characterization of their specific molecular target. Here, we describe a method for fast and efficient screening of venom and toxin activity using live-cell calcium imaging. We describe the use of Fura-2, a calcium indictor that changes its fluorescence properties in response to intracellular calcium elevations, to measure the activity of neurons from the dorsal root and trigeminal ganglia. Calcium imaging is an efficient technique for testing many of the venom's components on large numbers of neurons simultaneously. This technique offers a novel tool for low-cost and rapid characterization of functionally active toxins and their target receptors.

ACS Style

Yossi Maatuf; Avi Priel. High-Throughput Calcium Imaging Screen of Toxins' Function in Dissociated Sensory Neurons. Methods in Molecular Biology 2019, 2068, 275 -282.

AMA Style

Yossi Maatuf, Avi Priel. High-Throughput Calcium Imaging Screen of Toxins' Function in Dissociated Sensory Neurons. Methods in Molecular Biology. 2019; 2068 ():275-282.

Chicago/Turabian Style

Yossi Maatuf; Avi Priel. 2019. "High-Throughput Calcium Imaging Screen of Toxins' Function in Dissociated Sensory Neurons." Methods in Molecular Biology 2068, no. : 275-282.

Review
Published: 23 February 2019 in Toxins
Reads 0
Downloads 0

Chronic pain is a major medical issue which reduces the quality of life of millions and inflicts a significant burden on health authorities worldwide. Currently, management of chronic pain includes first-line pharmacological therapies that are inadequately effective, as in just a portion of patients pain relief is obtained. Furthermore, most analgesics in use produce severe or intolerable adverse effects that impose dose restrictions and reduce compliance. As the majority of analgesic agents act on the central nervous system (CNS), it is possible that blocking pain at its source by targeting nociceptors would prove more efficient with minimal CNS-related side effects. The development of such analgesics requires the identification of appropriate molecular targets and thorough understanding of their structural and functional features. To this end, plant and animal toxins can be employed as they affect ion channels with high potency and selectivity. Moreover, elucidation of the toxin-bound ion channel structure could generate pharmacophores for rational drug design while favorable safety and analgesic profiles could highlight toxins as leads or even as valuable therapeutic compounds themselves. Here, we discuss the use of plant and animal toxins in the characterization of peripherally expressed ion channels which are implicated in pain.

ACS Style

Yossi Maatuf; Matan Geron; Avi Priel. The Role of Toxins in the Pursuit for Novel Analgesics. Toxins 2019, 11, 131 .

AMA Style

Yossi Maatuf, Matan Geron, Avi Priel. The Role of Toxins in the Pursuit for Novel Analgesics. Toxins. 2019; 11 (2):131.

Chicago/Turabian Style

Yossi Maatuf; Matan Geron; Avi Priel. 2019. "The Role of Toxins in the Pursuit for Novel Analgesics." Toxins 11, no. 2: 131.

Journal article
Published: 21 November 2018 in Proceedings of the National Academy of Sciences
Reads 0
Downloads 0

Many neurotoxins inflict pain by targeting receptors expressed on nociceptors, such as the polymodal cationic channel TRPV1. The tarantula double-knot toxin (DkTx) is a peptide with an atypical bivalent structure, providing it with the unique capability to lock TRPV1 in its open state and evoke an irreversible channel activation. Here, we describe a distinct gating mechanism of DkTx-evoked TRPV1 activation. Interestingly, DkTx evokes significantly smaller TRPV1 macroscopic currents than capsaicin, with a significantly lower unitary conductance. Accordingly, while capsaicin evokes aversive behaviors in TRPV1-transgenic Caenorhabditis elegans, DkTx fails to evoke such response at physiological concentrations. To determine the structural feature(s) responsible for this phenomenon, we engineered and evaluated a series of mutated toxins and TRPV1 channels. We found that elongating the DkTx linker, which connects its two knots, increases channel conductance compared with currents elicited by the native toxin. Importantly, deletion of the TRPV1 pore turret, a stretch of amino acids protruding out of the channel’s outer pore region, is sufficient to produce both full conductance and aversive behaviors in response to DkTx. Interestingly, this deletion decreases the capsaicin-evoked channel activation. Taken together with structure modeling analysis, our results demonstrate that the TRPV1 pore turret restricts DkTx-mediated pore opening, probably through steric hindrance, limiting the current size and mitigating the evoked downstream physiological response. Overall, our findings reveal that DkTx and capsaicin elicit distinct TRPV1 gating mechanisms and subsequent pain responses. Our results also indicate that the TRPV1 pore turret regulates the mechanisms of channel gating and permeation.

ACS Style

Matan Geron; Rakesh Kumar; Wenchang Zhou; José D. Faraldo-Gómez; Valeria Vásquez; Avi Priel. TRPV1 pore turret dictates distinct DkTx and capsaicin gating. Proceedings of the National Academy of Sciences 2018, 115, E11837 -E11846.

AMA Style

Matan Geron, Rakesh Kumar, Wenchang Zhou, José D. Faraldo-Gómez, Valeria Vásquez, Avi Priel. TRPV1 pore turret dictates distinct DkTx and capsaicin gating. Proceedings of the National Academy of Sciences. 2018; 115 (50):E11837-E11846.

Chicago/Turabian Style

Matan Geron; Rakesh Kumar; Wenchang Zhou; José D. Faraldo-Gómez; Valeria Vásquez; Avi Priel. 2018. "TRPV1 pore turret dictates distinct DkTx and capsaicin gating." Proceedings of the National Academy of Sciences 115, no. 50: E11837-E11846.

Validation study
Published: 15 December 2017 in Diabetes
Reads 0
Downloads 0

Polymorphism in TCF7L2, a component of the canonical Wnt signaling pathway, has a strong association with β-cell dysfunction and type 2 diabetes through a yet to be defined mechanism. β-Cells rely on cells in their microenvironment, including pericytes, for their proper function. Here, we show that Tcf7l2 activity in pancreatic pericytes is required for β-cell function. Transgenic mice in which Tcf7l2 was selectively inactivated in their pancreatic pericytes exhibited impaired glucose tolerance due to compromised β-cell function and glucose-stimulated insulin secretion. Inactivation of pericytic Tcf7l2 was associated with impaired expression of genes required for β-cell function and maturity in isolated islets. In addition, we identified Tcf7l2-dependent pericytic expression of secreted factors shown to promote β-cell function, including BMP4. Finally, we show that exogenous BMP4 is sufficient to rescue the impaired glucose-stimulated insulin secretion of transgenic mice, pointing to a potential mechanism through which pericytic Tcf7l2 activity impacts β-cells. To conclude, we suggest that pancreatic pericytes produce secreted factors, including BMP4, in a Tcf7l2-dependent manner to support β-cell function. Our findings thus propose a potential cellular mechanism through which abnormal TCF7L2 activity predisposes individuals to diabetes, and implicate abnormalities in the islet microenvironment in this disease.

ACS Style

Lina Sakhneny; Eleonor Rachi; Alona Epshtein; Helen C. Guez; Shane Wald-Altman; Michal Lisnyansky; Laura Khalifa-Malka; Adina Hazan; Daria Baer; Avi Priel; Miguel Weil; Limor Landsman. Pancreatic Pericytes Support β-Cell Function in a Tcf7l2-Dependent Manner. Diabetes 2017, 67, 437 -447.

AMA Style

Lina Sakhneny, Eleonor Rachi, Alona Epshtein, Helen C. Guez, Shane Wald-Altman, Michal Lisnyansky, Laura Khalifa-Malka, Adina Hazan, Daria Baer, Avi Priel, Miguel Weil, Limor Landsman. Pancreatic Pericytes Support β-Cell Function in a Tcf7l2-Dependent Manner. Diabetes. 2017; 67 (3):437-447.

Chicago/Turabian Style

Lina Sakhneny; Eleonor Rachi; Alona Epshtein; Helen C. Guez; Shane Wald-Altman; Michal Lisnyansky; Laura Khalifa-Malka; Adina Hazan; Daria Baer; Avi Priel; Miguel Weil; Limor Landsman. 2017. "Pancreatic Pericytes Support β-Cell Function in a Tcf7l2-Dependent Manner." Diabetes 67, no. 3: 437-447.

Review
Published: 16 October 2017 in Toxins
Reads 0
Downloads 0

Beyond providing evolutionary advantages, venoms offer unique research tools, as they were developed to target functionally important proteins and pathways. As a key pain receptor in the nociceptive pathway, transient receptor potential vanilloid 1 (TRPV1) of the TRP superfamily has been shown to be a target for several toxins, as a way of producing pain to deter predators. Importantly, TRPV1 is involved in thermoregulation, inflammation, and acute nociception. As such, toxins provide tools to understand TRPV1 activation and modulation, a critical step in advancing pain research and the development of novel analgesics. Indeed, the phytotoxin capsaicin, which is the spicy chemical in chili peppers, was invaluable in the original cloning and characterization of TRPV1. The unique properties of each subsequently characterized toxin have continued to advance our understanding of functional, structural, and biophysical characteristics of TRPV1. By building on previous reviews, this work aims to provide a comprehensive summary of the advancements made in TRPV1 research in recent years by employing animal toxins, in particular DkTx, RhTx, BmP01, Echis coloratus toxins, APHCs and HCRG21. We examine each toxin’s functional aspects, behavioral effects, and structural features, all of which have contributed to our current knowledge of TRPV1. We additionally discuss the key features of TRPV1’s outer pore domain, which proves to be the target of the currently discussed toxins.

ACS Style

Matan Geron; Adina Hazan; Avi Priel. Animal Toxins Providing Insights into TRPV1 Activation Mechanism. Toxins 2017, 9, 326 .

AMA Style

Matan Geron, Adina Hazan, Avi Priel. Animal Toxins Providing Insights into TRPV1 Activation Mechanism. Toxins. 2017; 9 (10):326.

Chicago/Turabian Style

Matan Geron; Adina Hazan; Avi Priel. 2017. "Animal Toxins Providing Insights into TRPV1 Activation Mechanism." Toxins 9, no. 10: 326.

Journal article
Published: 13 October 2017 in Journal of the American Society of Nephrology
Reads 0
Downloads 0

Altered glucose reabsorption via the facilitative glucose transporter 2 (GLUT2) during diabetes may lead to renal proximal tubule cell (RPTC) injury, inflammation, and interstitial fibrosis. These pathologies are also triggered by activating the cannabinoid-1 receptor (CB1R), which contributes to the development of diabetic nephropathy (DN). However, the link between CB1R and GLUT2 remains to be determined. Here, we show that chronic peripheral CB1R blockade or genetically inactivating CB1Rs in the RPTCs ameliorated diabetes-induced renal structural and functional changes, kidney inflammation, and tubulointerstitial fibrosis in mice. Inhibition of CB1R also downregulated GLUT2 expression, affected the dynamic translocation of GLUT2 to the brush border membrane of RPTCs, and reduced glucose reabsorption. Thus, targeting peripheral CB1R or inhibiting GLUT2 dynamics in RPTCs has the potential to treat and ameliorate DN. These findings may support the rationale for the clinical testing of peripherally restricted CB1R antagonists or the development of novel renal-specific GLUT2 inhibitors against DN.

ACS Style

Liad Hinden; Shiran Udi; Adi Drori; Asaad Gammal; Alina Nemirovski; Rivka Hadar; Saja Baraghithy; Anna Permyakova; Matan Geron; Merav Cohen; Sabina Tsytkin-Kirschenzweig; Yael Riahi; Gil Leibowitz; Yaakov Nahmias; Avi Priel; Joseph Tam. Modulation of Renal GLUT2 by the Cannabinoid-1 Receptor: Implications for the Treatment of Diabetic Nephropathy. Journal of the American Society of Nephrology 2017, 29, 434 -448.

AMA Style

Liad Hinden, Shiran Udi, Adi Drori, Asaad Gammal, Alina Nemirovski, Rivka Hadar, Saja Baraghithy, Anna Permyakova, Matan Geron, Merav Cohen, Sabina Tsytkin-Kirschenzweig, Yael Riahi, Gil Leibowitz, Yaakov Nahmias, Avi Priel, Joseph Tam. Modulation of Renal GLUT2 by the Cannabinoid-1 Receptor: Implications for the Treatment of Diabetic Nephropathy. Journal of the American Society of Nephrology. 2017; 29 (2):434-448.

Chicago/Turabian Style

Liad Hinden; Shiran Udi; Adi Drori; Asaad Gammal; Alina Nemirovski; Rivka Hadar; Saja Baraghithy; Anna Permyakova; Matan Geron; Merav Cohen; Sabina Tsytkin-Kirschenzweig; Yael Riahi; Gil Leibowitz; Yaakov Nahmias; Avi Priel; Joseph Tam. 2017. "Modulation of Renal GLUT2 by the Cannabinoid-1 Receptor: Implications for the Treatment of Diabetic Nephropathy." Journal of the American Society of Nephrology 29, no. 2: 434-448.

Rapid communication
Published: 25 May 2017 in ACS Chemical Neuroscience
Reads 0
Downloads 0

TRPV1 is a prominent signal integrator of the pain system, known to be activated by vanilloids, a family of endogenous and exogenous pain-evoking molecules, through the vanilloid-binding site (VBS). The extensive preclinical profiling of small molecule inhibitors provides intriguing evidence that TRPV1 inhibition can be a useful therapeutic approach. However, the dissimilarity of chemical species that activate TRPV1 creates a major obstacle to understanding the molecular mechanism of pain induction, which is viewed as a pivotal trait of the somatosensory system. Here, we establish the existence of a unique family of synthetic agonists that interface with TRPV1 through the VBS, containing none of the molecular domains previously believed to be required for this interaction. The overarching value obtained from our inquiry is the novel advancement of the existing TRPV1 activation model. These findings uncover new potential in the area of pain treatment, providing a novel synthetic platform.

ACS Style

Yelena Mostinski; Gilad Noy; Rakesh Kumar; Dmitry Tsvelikhovsky; Avi Priel. Tricyclic Spirolactones as Modular TRPV1 Synthetic Agonists. ACS Chemical Neuroscience 2017, 8, 1688 -1696.

AMA Style

Yelena Mostinski, Gilad Noy, Rakesh Kumar, Dmitry Tsvelikhovsky, Avi Priel. Tricyclic Spirolactones as Modular TRPV1 Synthetic Agonists. ACS Chemical Neuroscience. 2017; 8 (8):1688-1696.

Chicago/Turabian Style

Yelena Mostinski; Gilad Noy; Rakesh Kumar; Dmitry Tsvelikhovsky; Avi Priel. 2017. "Tricyclic Spirolactones as Modular TRPV1 Synthetic Agonists." ACS Chemical Neuroscience 8, no. 8: 1688-1696.

Journal article
Published: 01 March 2017 in Biochimica et Biophysica Acta (BBA) - General Subjects
Reads 0
Downloads 0

Peptide and protein toxins are essential tools to dissect and probe the biology of their target receptors. Venoms target vital physiological processes to evoke pain. Snake venoms contain various factors with the ability to evoke, enhance and sustain pain sensation. While a number of venom-derived toxins were shown to directly target TRPV1 channels expressed on somatosensory nerve terminals to evoke pain response, such toxins were yet to be identified in snake venoms. We screened Echis coloratus saw-scaled viper venom's protein fractions isolated by reversed phase HPLC for their ability to activate TRPV1 channels. To this end, we employed heterologous systems to analyze TRPV1 and NGF pathways by imaging and electrophysiology, combined with molecular biology, biochemical, and pharmacological tools. We identified TRPV1 activating proteins in the venom of Echis coloratus that produce a channel-dependent increase in intracellular calcium and outwardly rectifying currents in neurons and heterologous systems. Interestingly, channel activation was not mediated by any of its known toxin binding sites. Moreover, although NGF neurotropic activity was detected in this venom, TRPV1 activation was independent of NGF receptors. Echis coloratus venom contains proteins with the ability to directly activate TRPV1. This activity is independent of the NGF pathway and is not mediated by known TRPV1 toxins' binding sites. Our results could facilitate the discovery of new toxins targeting TRPV1 to enhance current understanding of this receptor activation mechanism. Furthermore, the findings of this study provide insight into the mechanism through which snakes' venom elicit pain.

ACS Style

Matan Geron; Rakesh Kumar; Henry Matzner; Adi Lahiani; Galit Gincberg; Gadi Cohen; Philip Lazarovici; Avi Priel. Protein toxins of the Echis coloratus viper venom directly activate TRPV1. Biochimica et Biophysica Acta (BBA) - General Subjects 2017, 1861, 615 -623.

AMA Style

Matan Geron, Rakesh Kumar, Henry Matzner, Adi Lahiani, Galit Gincberg, Gadi Cohen, Philip Lazarovici, Avi Priel. Protein toxins of the Echis coloratus viper venom directly activate TRPV1. Biochimica et Biophysica Acta (BBA) - General Subjects. 2017; 1861 (3):615-623.

Chicago/Turabian Style

Matan Geron; Rakesh Kumar; Henry Matzner; Adi Lahiani; Galit Gincberg; Gadi Cohen; Philip Lazarovici; Avi Priel. 2017. "Protein toxins of the Echis coloratus viper venom directly activate TRPV1." Biochimica et Biophysica Acta (BBA) - General Subjects 1861, no. 3: 615-623.

Video audio media
Published: 17 February 2017 in Journal of Visualized Experiments
Reads 0
Downloads 0

Transfection, the delivery of foreign nucleic acids into a cell, is a powerful tool in protein research. Through this method, ion channels can be investigated through electrophysiological analysis, biochemical characterization, mutational studies, and their effects on cellular processes. Transient transfections offer a simple protocol in which the protein becomes available for analysis within a few hours to days. Although this method presents a relatively straightforward and time efficient protocol, one of the critical components is calibrating the expression of the gene of interest to physiological relevant levels or levels that are suitable for analysis. To this end, many different approaches that offer the ability to control the expression of the gene of interest have emerged. Several stable cell transfection protocols provide a way to permanently introduce a gene of interest into the cellular genome under the regulation of a tetracycline-controlled transcriptional activation. While this technique produces reliable expression levels, each gene of interest requires a few weeks of skilled work including calibration of a killing curve, selection of cell colonies, and overall more resources. Here we present a protocol that uses transient transfection of the Transient Receptor Potential cation channel subfamily V member 1 (TRPV1) gene in an inducible system as an efficient way to express a protein in a controlled manner which is essential in ion channel analysis. We demonstrate that using this technique, we are able to perform calcium imaging, whole cell, and single channel analysis with controlled channel levels required for each type of data collection with a single transfection. Overall, this provides a replicable technique that can be used to study ion channels structure and function.

ACS Style

Matan Geron; Adina Hazan; Avi Priel. Controllable Ion Channel Expression through Inducible Transient Transfection. Journal of Visualized Experiments 2017, 1 .

AMA Style

Matan Geron, Adina Hazan, Avi Priel. Controllable Ion Channel Expression through Inducible Transient Transfection. Journal of Visualized Experiments. 2017; (120):1.

Chicago/Turabian Style

Matan Geron; Adina Hazan; Avi Priel. 2017. "Controllable Ion Channel Expression through Inducible Transient Transfection." Journal of Visualized Experiments , no. 120: 1.

Research
Published: 16 December 2016 in The FASEB Journal
Reads 0
Downloads 0

Peripheral neuronal activation by inflammatory mediators is a multifaceted physiological response that involves a multitude of regulated cellular functions. One key pathway that has been shown to be involved in inflammatory pain is Gq/GPCR, whose activation by inflammatory mediators is followed by the regulated response of the cation channel transient receptor potential vanilloid 1 (TRPV1). However, the mechanism that underlies TRPV1 activation downstream of the Gq/GPCR pathway has yet to be fully defined. In this study, we employ pharmacological and molecular biology tools to dissect this activation mechanism via perforated-patch recordings and calcium imaging of both neurons and a heterologous system. We showed that TRPV1 activity downstream of Gq/GPCR activation only produced a subdued current, which was noticeably different from the robust current that is typical of TRPV1 activation by exogenous stimuli. Moreover, we specifically demonstrated that 2 pathways downstream of Gq/GPCR signaling, namely endovanilloid production by lipoxygenases and channel phosphorylation by PKC, converge on TRPV1 to evoke a tightly regulated response. Of importance, we show that only when both pathways are acting on TRPV1 is the inflammatory-mediated response achieved. We propose that the requirement of multiple signaling events allows subdued TRPV1 activation to evoke regulated neuronal response during inflammation.—Kumar R., Hazan, A., Geron, M., Steinberg, R., Livni, L., Matzner, H., Priel, A. Activation of transient receptor potential vanilloid 1 by lipoxygenase metabolites depends on PKC phosphorylation. FASEB J. 31, 1238–1247 (2017). www.fasebj.org

ACS Style

Rakesh Kumar; Adina Hazan; Matan Geron; Rebbeca Steinberg; Lital Livni; Henry Matzner; Avi Priel. Activation of transient receptor potential vanilloid 1 by lipoxygenase metabolites depends on PKC phosphorylation. The FASEB Journal 2016, 31, 1238 -1247.

AMA Style

Rakesh Kumar, Adina Hazan, Matan Geron, Rebbeca Steinberg, Lital Livni, Henry Matzner, Avi Priel. Activation of transient receptor potential vanilloid 1 by lipoxygenase metabolites depends on PKC phosphorylation. The FASEB Journal. 2016; 31 (3):1238-1247.

Chicago/Turabian Style

Rakesh Kumar; Adina Hazan; Matan Geron; Rebbeca Steinberg; Lital Livni; Henry Matzner; Avi Priel. 2016. "Activation of transient receptor potential vanilloid 1 by lipoxygenase metabolites depends on PKC phosphorylation." The FASEB Journal 31, no. 3: 1238-1247.

Journal article
Published: 01 June 2016 in Journal of Biological Chemistry
Reads 0
Downloads 0

Vanilloids are pain evoking molecules that serve as ligands of the heat and capsaicin receptor TRPV1. Binding of either endogenous or exogenous vanilloids evokes channel and subsequent neuronal activation, leading to pain sensation. Despite its pivotal physiological role, the molecular basis of TRPV1 activation and deactivation is not fully understood. The highly conserved tyrosine in position 511 (Y511) of the rat TRPV1 (rTRPV1) was the first residue to be identified as a necessary participant in the vanilloid-mediated response. rTRPV1 cryo-EM structures implicated rotation of this residue in the vanilloids bound state. Therefore, we hypothesize that rTRPV1 Y511 residue entraps vanilloids in their binding site, prolonging channel activity. To test our hypothesis, we generated an array of rTRPV1 mutants, containing the whole spectrum of Y511 substitutions, and tested their response to both exo- and endo-vanilloids. Our data show that only substitutions of Y511 to aromatic amino acids were able to mimic, albeit partially, the vanilloid-evoked activation pattern of the wt receptor. Although these substitutions reduced the channel sensitivity to vanilloids, a maximal open-channel lifetime could be achieved. Moreover, while their current activation rate remains intact, receptors with Y511 substitutions exhibited a faster current deactivation. Our findings therefore suggest that the duration of channel activity evoked by vanilloids is regulated by the interaction between Y511 and the agonist. To conclude, we suggest that Y511-mediated anchoring of vanilloids in their binding pocket is pivotal for TRPV1 activation and subsequent pain sensation.

ACS Style

Kumar Rakesh; Adina Hazan; Arijit Basu; Nomi Zalcman; Henry Matzner; Avi Priel. Tyrosine Residue in the TRPV1 Vanilloid Binding Pocket Regulates Deactivation Kinetics. Journal of Biological Chemistry 2016, 291, 13855 -13863.

AMA Style

Kumar Rakesh, Adina Hazan, Arijit Basu, Nomi Zalcman, Henry Matzner, Avi Priel. Tyrosine Residue in the TRPV1 Vanilloid Binding Pocket Regulates Deactivation Kinetics. Journal of Biological Chemistry. 2016; 291 (26):13855-13863.

Chicago/Turabian Style

Kumar Rakesh; Adina Hazan; Arijit Basu; Nomi Zalcman; Henry Matzner; Avi Priel. 2016. "Tyrosine Residue in the TRPV1 Vanilloid Binding Pocket Regulates Deactivation Kinetics." Journal of Biological Chemistry 291, no. 26: 13855-13863.

Journal article
Published: 01 February 2016 in Biophysical Journal
Reads 0
Downloads 0

Vanilloids are pain evoking molecules that serve as ligands of the ‘heat and capsaicin receptor’ TRPV1. It is expressed by primary afferent sensory neurons of the pain pathway, where it functions as a sensor of noxious heat (>42°C) and various noxious chemicals. TRPV1 channel is formed by the symmetrical arrangement of four identical subunits around a central ion-conducting pore. Binding of vanilloids evokes channel opening and subsequent neuronal activation, leading to pain sensation. Despite its pivotal physiological role, the molecular basis of TRPV1 activation by vanilloids is not fully understood.

ACS Style

Rakesh Kumar; Adina Hazan; Henry Matzner; Avi Priel. The Stoichiometry and Activation Mechanism of TRPV1 by Vanilloids. Biophysical Journal 2016, 110, 24a -25a.

AMA Style

Rakesh Kumar, Adina Hazan, Henry Matzner, Avi Priel. The Stoichiometry and Activation Mechanism of TRPV1 by Vanilloids. Biophysical Journal. 2016; 110 (3):24a-25a.

Chicago/Turabian Style

Rakesh Kumar; Adina Hazan; Henry Matzner; Avi Priel. 2016. "The Stoichiometry and Activation Mechanism of TRPV1 by Vanilloids." Biophysical Journal 110, no. 3: 24a-25a.

Journal article
Published: 21 July 2015 in Scientific Reports
Reads 0
Downloads 0

The receptor channel TRPV1 (Transient Receptor Potential Vanilloid 1) is expressed by primary afferent sensory neurons of the pain pathway, where it functions as a sensor of noxious heat and various chemicals, including eicosanoids, capsaicin, protons and peptide toxins. Comprised of four identical subunits that organize into a non-selective cationic permeable channel, this receptor has a variety of binding sites responsible for detecting their respective agonists. Although its physiological role as a chemosensor has been described in detail, the stoichiometry of TRPV1 activation by its different ligands remains unknown. Here, we combined the use of concatemeric constructs harboring mutated binding sites with patch-clamp recordings in order to determine the stoichiometry for TRPV1 activation through the vanilloid binding site and the outer-pore domain by capsaicin and protons, respectively. We show that, while a single capsaicin-bound subunit was sufficient to achieve a maximal open-channel lifetime, all four proton-binding sites were required. Thus, our results demonstrate a distinct stoichiometry of TRPV1 activation through two of its different agonist-binding domains.

ACS Style

Adina Hazan; Rakesh Kumar; Henry Matzner; Avi Priel. The pain receptor TRPV1 displays agonist-dependent activation stoichiometry. Scientific Reports 2015, 5, 12278 .

AMA Style

Adina Hazan, Rakesh Kumar, Henry Matzner, Avi Priel. The pain receptor TRPV1 displays agonist-dependent activation stoichiometry. Scientific Reports. 2015; 5 (1):12278.

Chicago/Turabian Style

Adina Hazan; Rakesh Kumar; Henry Matzner; Avi Priel. 2015. "The pain receptor TRPV1 displays agonist-dependent activation stoichiometry." Scientific Reports 5, no. 1: 12278.

Journal article
Published: 05 March 2015 in Cellular Signalling
Reads 0
Downloads 0

Direct interaction of α9β1 integrin with nerve growth factor (NGF) has been previously reported to induce pro-proliferative and pro-survival activities of non-neuronal cells. We investigated participation of p75NTR in α9β1 integrin-dependent cellular response to NGF stimulation. Using selective transfection of glioma cell lines with these receptors, we showed a strong, cation-independent association of α9 integrin subunit with p75NTR on the cellular membrane by selective immunoprecipitation experiments. The presence of the α9/p75NTR complex increases NGF-dependent cell adhesion, proliferation and migration. Other integrin subunits including β1 were not found in complex with p75NTR. FRET analysis indicated that p75NTR and α9 integrin subunit are not closely associated through their cytoplasmic domains, most probably because of the molecular interference with other cytoplasmic proteins such as paxillin. Interaction of α9β1 integrin with another ligand, VCAM-1 was not modulated by the p75NTR. α9/p75NTR complex elevated NGF-dependent activation of MAPK Erk1/2 arty for integrin that may create active complexes with other types of receptors belonging to the TNF superfamily.

ACS Style

Erin M. Ventresca; Shimon Lecht; Piotr Jakubowski; Rachel A. Chiaverelli; Michael Weaver; Luis Del Valle; Keren Ettinger; Galit Gincberg; Avi Priel; Alex Braiman; Philip Lazarovici; Peter I. Lelkes; Cezary Marcinkiewicz. Association of p75NTR and α9β1 integrin modulates NGF-dependent cellular responses. Cellular Signalling 2015, 27, 1225 -1236.

AMA Style

Erin M. Ventresca, Shimon Lecht, Piotr Jakubowski, Rachel A. Chiaverelli, Michael Weaver, Luis Del Valle, Keren Ettinger, Galit Gincberg, Avi Priel, Alex Braiman, Philip Lazarovici, Peter I. Lelkes, Cezary Marcinkiewicz. Association of p75NTR and α9β1 integrin modulates NGF-dependent cellular responses. Cellular Signalling. 2015; 27 (6):1225-1236.

Chicago/Turabian Style

Erin M. Ventresca; Shimon Lecht; Piotr Jakubowski; Rachel A. Chiaverelli; Michael Weaver; Luis Del Valle; Keren Ettinger; Galit Gincberg; Avi Priel; Alex Braiman; Philip Lazarovici; Peter I. Lelkes; Cezary Marcinkiewicz. 2015. "Association of p75NTR and α9β1 integrin modulates NGF-dependent cellular responses." Cellular Signalling 27, no. 6: 1225-1236.

Journal article
Published: 28 May 2010 in Cell
Reads 0
Downloads 0

Summary Toxins have evolved to target regions of membrane ion channels that underlie ligand binding, gating, or ion permeation, and have thus served as invaluable tools for probing channel structure and function. Here, we describe a peptide toxin from the Earth Tiger tarantula that selectively and irreversibly activates the capsaicin- and heat-sensitive channel, TRPV1. This high-avidity interaction derives from a unique tandem repeat structure of the toxin that endows it with an antibody-like bivalency. The "double-knot" toxin traps TRPV1 in the open state by interacting with residues in the presumptive pore-forming region of the channel, highlighting the importance of conformational changes in the outer pore region of TRP channels during activation.

ACS Style

Christopher J. Bohlen; Avi Priel; Sharleen Zhou; David King; Jan Siemens; David Julius. A Bivalent Tarantula Toxin Activates the Capsaicin Receptor, TRPV1, by Targeting the Outer Pore Domain. Cell 2010, 141, 834 -845.

AMA Style

Christopher J. Bohlen, Avi Priel, Sharleen Zhou, David King, Jan Siemens, David Julius. A Bivalent Tarantula Toxin Activates the Capsaicin Receptor, TRPV1, by Targeting the Outer Pore Domain. Cell. 2010; 141 (5):834-845.

Chicago/Turabian Style

Christopher J. Bohlen; Avi Priel; Sharleen Zhou; David King; Jan Siemens; David Julius. 2010. "A Bivalent Tarantula Toxin Activates the Capsaicin Receptor, TRPV1, by Targeting the Outer Pore Domain." Cell 141, no. 5: 834-845.

Comparative study
Published: 01 June 2007 in Biophysical Journal
Reads 0
Downloads 0

Patch-clamp recording has revolutionized the study of ion channels, transporters, and the electrical activity of small cells. Vital to this method is formation of a tight seal between glass recording pipette and cell membrane. To better understand seal formation and improve practical application of this technique, we examine the effects of divalent ions, protons, ionic strength, and membrane proteins on adhesion of membrane to glass and on seal resistance using both patch-clamp recording and atomic force microscopy. We find that H+, Ca2+, and Mg2+ increase adhesion force between glass and membrane (lipid and cellular), decrease the time required to form a tight seal, and increase seal resistance. In the absence of H+ (10−10M) and divalent cations (<10−8M), adhesion forces are greatly reduced and tight seals are not formed. H+ (10−7M) promotes seal formation in the absence of divalent cations. A positive correlation between adhesion force and seal formation indicates that high resistance seals are associated with increased adhesion between membrane and glass. A similar ionic dependence of the adhesion of lipid membranes and cell membranes to glass indicates that lipid membranes without proteins are sufficient for the action of ions on adhesion

ACS Style

Avi Priel; Ziv Gil; Vincent T. Moy; Karl L. Magleby; Shai D. Silberberg. Ionic Requirements for Membrane-Glass Adhesion and Giga Seal Formation in Patch-Clamp Recording. Biophysical Journal 2007, 92, 3893 -3900.

AMA Style

Avi Priel, Ziv Gil, Vincent T. Moy, Karl L. Magleby, Shai D. Silberberg. Ionic Requirements for Membrane-Glass Adhesion and Giga Seal Formation in Patch-Clamp Recording. Biophysical Journal. 2007; 92 (11):3893-3900.

Chicago/Turabian Style

Avi Priel; Ziv Gil; Vincent T. Moy; Karl L. Magleby; Shai D. Silberberg. 2007. "Ionic Requirements for Membrane-Glass Adhesion and Giga Seal Formation in Patch-Clamp Recording." Biophysical Journal 92, no. 11: 3893-3900.

Journal article
Published: 21 December 2006 in Neuron
Reads 0
Downloads 0

10 páginas, 7 figuras.A prominent feature of ionotropic glutamate receptors from the AMPA and kainate subtypes is their profound desensitization in response to glutamate—a process thought to protect the neuron from overexcitation. In AMPA receptors, it is well established that desensitization results from rearrangements of the interface formed between agonist-binding domains of adjacent subunits; however, it is unclear how this mechanism applies to kainate receptors. Here we show that stabilization of the binding domain dimer by the generation of intermolecular disulfide bonds apparently blocked desensitization of the kainate receptor GluR6. This result establishes a common desensitization mechanism in both AMPA and kainate receptors. Surprisingly, however, surface expression of these nondesensitizing mutants was drastically reduced and did not depend on channel activity. Therefore, in addition to its role at the synapse, we now propose an intracellular role for desensitization in controlling maturation and trafficking of glutamate receptors.This work was supported by grants to Y.S.-B. from the Israel Science\ud Foundation (grant 561/03) and the European Commission (EUSynapse\ud project; contract LSHM-CT-2005-019055) and to J.L. by the\ud Spanish Ministry of Education and Science (grant BFU2006-\ud 07138). A.P. is a recipient of the David Kline prize of excellence by\ud the Canadian Friends of the Hebrew University. S.S. is an I3P Program\ud CSIC Research Fellow.Peer reviewe

ACS Style

Avi Priel; Sanja Selak; Juan Lerma; Yael Stern-Bach. Block of Kainate Receptor Desensitization Uncovers a Key Trafficking Checkpoint. Neuron 2006, 52, 1037 -1046.

AMA Style

Avi Priel, Sanja Selak, Juan Lerma, Yael Stern-Bach. Block of Kainate Receptor Desensitization Uncovers a Key Trafficking Checkpoint. Neuron. 2006; 52 (6):1037-1046.

Chicago/Turabian Style

Avi Priel; Sanja Selak; Juan Lerma; Yael Stern-Bach. 2006. "Block of Kainate Receptor Desensitization Uncovers a Key Trafficking Checkpoint." Neuron 52, no. 6: 1037-1046.