Journal club 2015.07.10.

HTR7 Mediates Serotonergic Acute and Chronic Itch

HTR7 Mediates Serotonergic Acute and Chronic Itch

Takeshi Morita,1,2,7 Shannan P. McClain,1,7 Lyn M. Batia,1 Maurizio Pellegrino,1 Sarah R. Wilson,1,2 Michael A. Kienzler,3 Kyle Lyman,3 Anne Sofie Braun Olsen,3 Justin F. Wong,1 Cheryl L. Stucky,4 Rachel B. Brem,5,6,* and Diana M. Bautista1,2,* 1Department of Molecular & Cell Biology, 142 Life Sciences Addition, University of California, Berkeley, Berkeley, CA 94720-3200, USA 2Helen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, CA 94720, USA
3Neurobiology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA
4Departments of Cell Biology, Neurobiology and Anatomy, Medical College of Wisconsin, Milwaukee, WI 53226, USA 5Buck Institute for Research on Aging, 8001 Redwood Boulevard, Novato, CA 94945, USA
6Department of Plant and Microbial Biology, University of California, Berkeley, Berkeley, CA 94720, USA
7Co-first author
*Correspondence: rbrem@buckinstitute.org (R.B.B.), dbautista@berkeley.edu (D.M.B.) http://dx.doi.org/10.1016/j.neuron.2015.05.044

SUMMARY
Chronic itch is a prevalent and debilitating condition for which few effective therapies are available. We harnessed the natural variation across genetically distinct mouse strains to identify transcripts co-re- gulated with itch behavior. This survey led to the discovery of the serotonin receptor HTR7 as a key mediator of serotonergic itch. Activation of HTR7 promoted opening of the ion channel TRPA1, which in turn triggered itch behaviors. In addition, acute itch triggered by serotonin or a selective serotonin reuptake inhibitor required both HTR7 and TRPA1. Aberrant serotonin signaling has long been linked to a variety of human chronic itch conditions, in- cluding atopic dermatitis. In a mouse model of atopic dermatitis, mice lacking HTR7 or TRPA1 dis- played reduced scratching and skin lesion severity. These data highlight a role for HTR7 in acute and chronic itch and suggest that HTR7 antagonists may be useful for treating a variety of pathological itch conditions.

Journal club 2015.07.10. Read More »

Journal Club 2015.06.12.

RESEARCH ARTICLE

Five hTRPA1 Agonists Found in Indigenous Korean Mint, Agastache rugosa

journal.pone.0127060

Hana Moon1☯, Min Jung Kim1☯, Hee Jin Son1, Hae-Jin Kweon2, Jung Tae Kim1, Yiseul Kim1, Jaewon Shim1, Byung-Chang Suh2, Mee-Ra Rhyu1*
1 Research Group of Food Functionality, Korea Food Research Institute, Bundang-gu, Sungnam-si, Gyeonggi-do, Republic of Korea, 2 Department of Brain Science, Daegu Gyeongbuk Institute of Science and Technology (DGIST), Daegu, Republic of Korea
☯ These authors contributed equally to this work. * mrrhyu@kfri.re.kr

Abstract
Transient receptor potential ankyrin1 (TRPA1) and transient receptor potential vanilloid 1 (TRPV1) are members of the TRP superfamily of structurally related, nonselective cation channels and mediators of several signaling pathways. Previously, we identified methyl syr- ingate as an hTRPA1 agonist with efficacy against gastric emptying. The aim of this study was to find hTRPA1 and/or hTRPV1 activators in Agastache rugosa (Fisch. et Meyer) O. Kuntze (A.rugosa), commonly known as Korean mint to improve hTRPA1-related phenome- na. An extract of the stem and leaves of A.rugosa (Labiatae) selectively activated hTRPA1 and hTRPV1. We next investigated the effects of commercially available compounds found in A.rugosa (acacetin, 4-allylanisole, p-anisaldehyde, apigenin 7-glucoside, L-carveol, β- caryophyllene, trans-p-methoxycinnamaldehyde, methyl eugenol, pachypodol, and ros- marinic acid) on cultured hTRPA1- and hTRPV1-expressing cells. Of the ten compounds, L-carveol, trans-p-methoxycinnamaldehyde, methyl eugenol, 4-allylanisole, and p-anisal- dehyde selectively activated hTRPA1, with EC50 values of 189.1±26.8, 29.8±14.9, 160.2 ±21.9, 1535±315.7, and 546.5±73.0 μM, respectively. The activities of these compounds were effectively inhibited by the hTRPA1 antagonists, ruthenium red and HC-030031. Al- though the five active compounds showed weaker calcium responses than allyl isothiocya- nate (EC50=7.2±1.4 μM), our results suggest that these compounds from the stem and leaves of A.rugosa are specific and selective agonists of hTRPA1.

Journal Club 2015.06.12. Read More »

Journal Club 2015.4.17

Structure of the TRPA1 ion channel suggests regulatory mechanisms

Candice E. Paulsen1*, Jean-Paul Armache2*, Yuan Gao1,2, Yifan Cheng2 & David Julius1

doi:10.1038/nature14367 nature14367

The TRPA1 ion channel (also known as the wasabi receptor) is a detector of noxious chemical agents encountered in our environment or produced endogenously during tissue injury or drug metabolism. These include a broad class of electrophiles that activate the channel through covalent protein modification. TRPA1 antagonists hold potential for treating neurogenic inflammatory conditions provoked or exacerbated by irritant exposure. Despite compelling reasons to understand TRPA1 function, structural mechanisms underlying channel regulation remain obscure. Here we use single-particle electron cryo- microscopy to determine the structure of full-length human TRPA1 to 4 A ̊ resolution in the presence of pharmacophores, including a potent antagonist. Several unexpected features are revealed, including an extensive coiled-coil assembly domain stabilized by polyphosphate co-factors and a highly integrated nexus that converges on an unpredicted transient receptor potential (TRP)-like allosteric domain. These findings provide new insights into the mechanisms of TRPA1 regulation, and establish a blueprint for structure-based design of analgesic and anti-inflammatory agents.

Journal Club 2015.4.17 Read More »

journal club 2015-03-06

5-HT3 receptors antagonists reduce serotonin-induced scratching in mice

Running title: 5-HT3 receptors mediate serotonin-induced scratching

fcp12112

Sattar Ostadhadi a,b, Nastaran Kordjazy a,b, Arya Haj-Mirzaian a,b, Parvin Mansouri c, Ahmad Reza Dehpour a,b*

a Experimental Medicine Research Center, Tehran University of Medical Sciences, Tehran, Iran

b Department of Pharmacology, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran

c Skin and Stem cell Research Center, Tehran University of Medical Sciences, Tehran, Iran

* Corresponding author:

Prof. Ahmad Reza Dehpour, PharmD, PhD, Department of Pharmacology, School of Medicine, Tehran University of Medical Sciences, P.O. Box 13145-784, Tehran, Iran. Tel: +98 21 88973652, fax: + 98 21 66402569, e-mail: dehpoura@sina.tums.ac.ir

 

ABSTRACT

Serotonin (5-hydroxytryptamine, 5-HT) acts as a pruritogen in humans and animals, but the mechanisms of action through that serotonin induces itch-response have not been extensively discovered. In our study, we attempted to investigate the role of 5-HT3 receptors in scratching behavior due to intradermal serotonin injection. Intradermal injection of serotonin (14.1–235 nmol/site) into the nape of the neck of mice was performed to elicit itch. Scratching behavior was evaluated by measuring the number of bouts during 60 minutes after injection. We evaluated the effect of intraperitoneal pretreatment with ondansetron and tropisetron (0.1, 0.3, and 1 mg/kg) on itch induced by serotonin. Also, intradermal ondansetron and tropisetron at doses 50, 100, and 200 nmol/site were concurrently administrated with serotonin. Serotonin produced a significant enhancement in scratching at dose 141 nmol/site. Concurrent administration of ondansetron (50, 100, and 200 nmol/site) and tropisetron (100 and 200 nmol/site) with serotonin reduced scratching activity compared to the animals that only received serotonin. Also, pretreatment with intraperitoneal ondansetron and tropisetron (0.3 and 1 mg/kg) 30 min before serotonin attenuated the itch response. We showed that the scratching induced by intradermal serotonin is mediated by 5-HT3 receptors subtype. It can be concluded that 5-HT3 may play a role in mediating serotonin-associated itch responses, and we introduce 5-HT3 receptors as possible targets for antipruritic agents.

Key words: Scratching; Serotonin (5-HT); 5-HT3 antagonists; Mice

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journal club 2015.01.23.

Protein kinase Cdelta mediates histamine-evoked itch and responses in pruriceptors

Molecular Pain 2015, 11:1 doi:10.1186/1744-8069-11-1

Manouela V Valtcheva (valtchevam@wusm.wustl.edu) Steve Davidson (sdavidson@wustl.edu) Chengshui Zhao (zhaoc@anest.wustl.edu) Michael Leitges (michael.leitges@biotek.uio.no) Robert W Gereau IV (gereaur@wustl.edu)

1744-8069-11-1

Abstract Background

Itch-producing compounds stimulate receptors expressed on small diameter fibers that innervate the skin. Many of the currently known pruritogen receptors are Gq-Protein Coupled Receptors (GqPCR), which activate Protein Kinase C (PKC). Specific isoforms of PKC have been previously shown to perform selective functions; however, the roles of PKC isoforms in regulating itch remain unclear. In this study, we investigated the novel PKC isoform PKCδ as an intracellular modulator of itch signaling in response to histamine and the non- histaminergic pruritogens chloroquine and β-alanine.

Results

Behavioral experiments indicate that PKCδ knock-out (KO) mice have a 40% reduction in histamine-induced scratching when compared to their wild type littermates. On the other hand, there were no differences between the two groups in scratching induced by the MRGPR agonists chloroquine or β-alanine. PKCδ was present in small diameter dorsal root ganglion (DRG) neurons. Of PKCδ-expressing neurons, 55% also stained for the non-peptidergic marker IB4, while a smaller percentage (15%) expressed the peptidergic marker CGRP. Twenty-nine percent of PKCδ-expressing neurons also expressed TRPV1. Calcium imaging studies of acutely dissociated DRG neurons from PKCδ-KO mice show a 40% reduction in the total number of neurons responsive to histamine. In contrast, there was no difference in the number of capsaicin-responsive neurons between KO and WT animals. Acute pharmacological inhibition of PKCδ with an isoform-specific peptide inhibitor (δV1-1) also significantly reduced the number of histamine-responsive sensory neurons.

Conclusions

Our findings indicate that PKCδ plays a role in mediating histamine-induced itch, but may be dispensable for chloroquine- and β-alanine-induced itch.

journal club 2015.01.23. Read More »

journal club – 2014.12.15.

A natural dye, Niram improves atopic dermatitis through down-regulation of TSLP

Na-Ra Hana, Jin-Young Parkb, Jae-Bum Jangb, Hyun-Ja Jeongc,∗, Hyung-Min Kima,∗∗

a Department of Pharmacology, College of Korean Medicine, Kyung Hee University, 1 Hoegi-dong, Dongdaemun-gu, Seoul 130-701, Republic of Korea
b Regional Innovation Center and Inflammatory Disease Research Center, Hoseo University, 165, Sechul-ri, Baebang-myun, Asan, Chungnam 336-795, Republic of Korea

c Department of Food Technology, Biochip Research Center, and Inflammatory Disease Research Center, Hoseo University, 165, Sechul-ri, Baebang-myun, Asan, Chungnam 336-795,Republic of Korea

1-s2.0-S1382668914002476-main

abstract

Naju Jjok (Polygonum tinctorium Lour.) has been known to treat skin diseases in traditional Korean medicine. A natural textile dye, Niram made from Naju Jjok has traditionally been used to dye clothes. Thymic stromal lymphopoietin (TSLP) plays an important role in the development of atopic dermatitis (AD). Thus, we investigated that Niram might amelio- rate AD through regulation of TSLP. Niram significantly inhibited the levels of TSLP through blockade of caspase-1/receptor-interacting protein 2 pathway in stimulated mast cells. Fur- ther, Niram ameliorated clinical symptoms in AD mouse. Niram significantly inhibited the infiltration of inflammatory cells in lesional skin. The levels of TSLP, caspase-1, IL-4, and IL-6 were inhibited in lesional skin applied topically with Niram. Niram significantly inhibited the serum levels of IgE and histamine in AD mouse. Finally, Niram significantly inhibited the levels of TSLP in polyriboinosinic polyribocytidylic acid-stimulated human keratinocyte HaCaT cells. These results establish Niram as a functional dye embracing the aspects of not only a traditional use but also a pharmacological effect.

© 2014 Elsevier B.V. All rights reserved.

Keywords : Niram, TSLP, Caspase-1, Mast cell, Atopic dermatitis, NC/Nga mice

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journal club – 14.10.27.

Propofol-induced pain sensation involves multiple mechanisms in sensory neurons

Rei Nishimoto & Makiko Kashio & Makoto Tominaga

Propofol

Received: 18 June 2014 / Revised: 22 September 2014 / Accepted: 25 September 2014 # Springer-Verlag Berlin Heidelberg 2014

Abstract Propofol, a commonly used intravenous anesthetic agent, is known to at times cause pain sensation upon injection in humans. However, the molecular mechanisms underlying this effect are not fully understood. Although propofol was reported to activate human transient receptor potential ankyrin 1 (TRPA1) in this regard, its action on human TRP vanilloid 1 (TRPV1), another nociceptive receptor, is unknown. Further- more, whether propofol activates TRPV1 in rodents is con- troversial. Here, we show that propofol activates human and mouse TRPA1. In contrast, we did not observe propofol- evoked human TRPV1 activation, while the ability of propofol to activate mouse TRPV1 was very small. We also found that propofol caused increases in intracellular Ca2+ concentrations in a considerable portion of dorsal root gangli- on (DRG) cells from mice lacking both TRPV1 and TRPA1, indicating the existence of TRPV1- and TRPA1-independent mechanisms for propofol action. In addition, propofol pro- duced action potential generation in a type A γ-amino butyric acid (GABAA) receptor-dependent manner. Finally, we found that both T-type and L-type Ca2+ channels are activated downstream of GABAA receptor activation by propofol. Thus, we conclude that propofol may cause pain sensation through multiple mechanisms involving not only TRPV1 and TRPA1 but also voltage-gated channels downstream of GABAA receptor activation.

Keywords Propofol . TRPV1 . TRPA1 . Voltage-gated Ca2+ channel . GABAA receptor

journal club – 14.10.27. Read More »

journal club 20-10-2014

Extracellular MicroRNAs Activate Nociceptor Neurons to Elicit Pain via TLR7 and TRPA1

Chul-Kyu Park,1,2 Zhen-Zhong Xu,1,2 Temugin Berta,1,2 Qingjian Han,1 Gang Chen,1 Xing-Jun Liu,1 and Ru-Rong Ji1,* 1Pain Signaling and Plasticity Laboratory, Departments of Anesthesiology and Neurobiology, Duke University Medical Center, Durham, NC 27710, USA
2Co-first authors

*Correspondence: ru-rong.ji@duke.edu http://dx.doi.org/10.1016/j.neuron.2014.02.011 Extracellular MicroRNAs Activate Nociceptor neurins to elicit pain via TLR7 and TRPV1

Intracellular microRNAs (miRNAs) are key regulators of gene expression. The role of extracellular miRNAs in neuronal activation and sensory behaviors are unknown. Here we report an unconventional role of extracellular miRNAs for rapid excitation of nocicep- tor neurons via toll-like receptor-7 (TLR7) and its coupling to TRPA1 ion channel. miRNA-let-7b induces rapid inward currents and action potentials in dorsal root ganglion (DRG) neurons. These re- sponses require the GUUGUGU motif, only occur in neurons coexpressing TLR7 and TRPA1, and are abolished in mice lacking Tlr7 or Trpa1. Furthermore, let-7b induces TLR7/TRPA1-dependent single-chan- nel activities in DRG neurons and HEK293 cells over- expressing TLR7/TRPA1. Intraplantar injection of let-7b elicits rapid spontaneous pain via TLR7 and TRPA1. Finally, let-7b can be released from DRG neurons by neuronal activation, and let-7b inhibitor reduces formalin-induced TRPA1 currents and spon- taneous pain. Thus, secreted extracellular miRNAs may serve as novel pain mediators via activating TLR7/TRPA1 in nociceptor neurons.

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Journal Club 2014. 09. 29.

The Bile Acid Receptor TGR5 Activates the TRPA1 Channel to Induce Itch in Mice Short Title: Bile Acid Evoked Itch

The Bile Acid Receptor TGR5 Activates the TRPA1 Channel to Induce Itch in Mice

TinaMarie Lieu, Gihan Jayaweera, Peishen Zhao, Daniel P. Poole, Dane Jensen, Megan Grace, Peter McIntyre, Romke Bron, Yvette M. Wilson, Matteus Krappitz, Silke Haerteis, Christoph Korbmacher, Martin S. Steinhoff, Romina Nassini, Serena Materazzi, Pierangelo Geppetti, Carlos U. Corvera, Nigel W. Bunnett

Abstract
Background & Aims: Patients with cholestatic disease have increased systemic concentrations of bile acids (BAs) and profound pruritus. The G protein-coupled BA receptor 1 TGR5 (encoded by GPBAR1) is expressed by primary sensory neurons; its activation induces neuronal hyperexcitability and scratching, by unknown mechanisms. We investigated whether the transient receptor potential ankyrin 1 (TRPA1) is involved in BA-evoked, TGR5-dependent pruritus in mice.

Methods: Co-expression of TGR5 and TRPA1 in cutaneous afferent neurons isolated from mice

was analyzed by immunofluorescence, in situ hybridization, and single-cell PCR. TGR5-induced

activation of TRPA1 was studied in in HEK293 cells, Xenopus laevis oocytes, and primary 2+

sensory neurons by measuring Ca signals. The contribution of TRPA1 to TGR5-induced release of pruritogenic neuropeptides, activation of spinal neurons, and scratching behavior were studied using TRPA1 antagonists or Trpa1–/– mice.

Results: TGR5 and TRPA1 protein and mRNA were expressed by cutaneous afferent neurons. In

HEK cells, oocytes, and neurons co-expressing TGR5 and TRPA1, BAs caused TGR5-dependent

activation and sensitization of TRPA1 by mechanisms that required Gβγ, protein kinase C and 2+

Ca . Antagonists or deletion of TRPA1 prevented BA-stimulated release of the pruritogenic neuropeptides gastrin-releasing peptide and atrial natriuretic peptide B in the spinal cord. Disruption of Trpa1 in mice blocked BA-induced expression of Fos in spinal neurons and prevented BA-stimulated scratching. Spontaneous scratching was exacerbated in transgenic mice that overexpressed TRG5. Administration of a TRPA1 antagonist or the BA sequestrant colestipol, which lowered circulating levels of BAs, prevented exacerbated spontaneous scratching in TGR5 overexpressing mice.

Conclusions: BAs induce pruritus in mice by co-activation of TGR5 and TRPA1. Antagonists of TGR5 and TRPA1, or inhibitors of the signaling mechanism by which TGR5 activates TRPA1, might be developed for treatment of cholestatic pruritus.

KEYWORDS: liver, mouse model, itching, signal transduction

Journal Club 2014. 09. 29. Read More »

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