Journal club 2014.07.25.

Activation of TRPV1 mediates thymic stromal lymphopoietin release via the Ca2+/NFAT pathway in airway epithelial cells

Xinying Jia a, Hong Zhang a, Xu Cao b,⇑, Yuxin Yin a, Bo Zhang a,⇑ a Department of Pathology, Peking University Health Science Center, 100191 Beijing, China

b Department of Neurology, Peking University Health Science Center, 100191 Beijing, China

1-s2.0-S0014579314004773-main-2 S0014579314004773-fx1.jpg

1-s2.0-S0014579314004773-main-2

Abstract

The airway epithelium is exposed to a range of irritants in the environment that are known to trig- ger inflammatory response such as asthma. Transient receptor potential vanilloid 1 (TRPV1) is a Ca2+-permeable cation channel critical for detecting noxious stimuli by sensory neurons. Recently increasing evidence suggests TRPV1 is also crucially involved in the pathophysiology of asthma on airway epithelium in human. Here we report that in airway epithelial cells TRPV1 activation potently induces allergic cytokine thymic stromal lymphopoietin (TSLP) release. TSLP induction by protease-activated receptor (PAR)-2 activation is also partially mediated by TRPV1 channels.

Ó 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

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Journal club 2014-06-27

Ligand determinants of fatty acid activation of the pronociceptive ion channel TRPA1

peerj-248

William John Redmond1, Liuqiong Gu2, Maxime Camo1, Peter McIntyre2,3 and Mark Connor1

  1. 1  Australian School of Advanced Medicine, Macquarie University, NSW, Australia
  2. 2  Department of Pharmacology, University of Melbourne, Parkville, Victoria, Australia
  3. 3  Health Innovations Research Institute and School of Medical Sciences, RMIT University, Melbourne,

    Victoria, Australia

ABSTRACT

Background and purpose. Arachidonic acid (AA) and its derivatives are important modulators of cellular signalling. The transient receptor potential cation channel subfamily A, member 1 (TRPA1) is a cation channel with important functions in mediating cellular responses to noxious stimuli and inflammation. There is limited information about the interactions between AA itself and TRPA1, so we investigated the effects of AA and key ethanolamide and amino acid/neurotransmitter deriva- tives of AA on hTRPA1.

Experimental approach. HEK 293 cells expressing hTRPA1 were studied by mea- suring changes in intracellular calcium ([Ca]i) with a fluorescent dye and by stan- dard whole cell patch clamp recordings.
Key results. AA (30 μM) increased fluorescence in hTRPA1 expressing cells by 370% (notional EC50 13 μM). The covalent TRPA1 agonist cinnamaldehyde

(300 μM) increased fluorescence by 430% (EC50, 11 μM). Anandamide (230%) and N -arachidonoyl tyrosine (170%) substantially activated hTRPA1 at 30 μM, how- ever, N -arachidonoyl conjugates of glycine and taurine were less effective while
N -acyl conjugates of 5-HT did not affect hTRPA1. Changing the acyl chain length or the number and position of double bonds reduced fatty acid efficacy at hTRPA1. Mutant hTRPA1 (Cys621, Cys641 and Cys665 changed to Ser) could be activated by AA (100 μM, 40% of wild type) but not by cinnamaldehyde (300 μM). Conclusions and implications. AA is a more potent activator of TRPA1 than its ethanolamide or amino acid/neurotransmitter derivatives and acts via a mechanism distinct from that of cinnamaldehyde, further underscoring the likelyhood of multi- ple pharmacologically exploitable sites on hTRPA1.

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Journal Club 2014-04-11

BEHAVIORAL MODEL OF ITCH, ALLOKNESIS, PAIN AND ALLODYNIA IN THE LOWER HINDLIMB AND CORRELATIVE RESPONSES
OF LUMBAR DORSAL HORN NEURONS IN THE MOUSE

T. AKIYAMA, M. NAGAMINE, M. I. CARSTENS AND E. CARSTENS *

University of California, Davis, Department of Neurobiology, Physiology & Behavior, 1 Shields Avenue, Davis, CA 95616, USA

1-s2.0-S0306452214001018-main

Abstract—We have further developed a behavioral model of itch and pain in the lower hindlimb (calf) originally reported by LaMotte et al. (2011) that allows comparisons with responses of lumbar dorsal horn neurons to pruritic and noxious stimuli. Intradermal (id) microinjection of the prurit- ogens histamine, SLIGRL-NH2 (agonist of PAR-2 and MrgprC11) and chloroquine (agonist of MrgprA3) into the calf of the lower limb elicited significant biting and a small amount of licking directed to the injection site, over a 30- min time course. Following id injection of histamine, low-threshold mechanical stimuli reliably elicited discrete episodes of biting (alloknesis) over a longer time course; significantly less alloknesis was observed following id injec- tion of SLIGRL-NH2. Capsaicin injections elicited licking but little biting. Following id injection of capsaicin, low-thresh- old mechanical stimuli elicited discrete hindlimb flinches (allodynia) over a prolonged (>2 h) time course. In single- unit recordings from superficial lumbar dorsal horn neurons, low-threshold mechanically evoked responses were significantly enhanced, accompanied by receptive field expansion, following id injection of histamine in histamine- responsive neurons. This was not observed in histamine- insensitive neurons, or following id injection of saline or SLIGRL-NH2, regardless of whether the latter activated the neuron or not. These results suggest that itch-responsive neurons are selectively sensitized by histamine but not SLI- GRL-NH2 to account for alloknesis. The presently described ‘‘calf’’ model appears to distinguish between itch- and pain- related behavioral responses, and provides a basis to inves- tigate lumbar spinal neural mechanisms underlying itch, alloknesis, pain and allodynia. Ó 2014 Published by Elsevier Ltd. on behalf of IBRO.

Key words: itch, alloknesis, pain, allodynia, scratching, dor- sal horn neuron.

Journal Club 2014-04-11 Read More »

Journal Club 2014-03-28

TRPV3 is a calcium-permeable temperature-sensitive cation channel

12077604-Nature-2002-TRPV3 is a calcium-permeable temperature-sensitive

Haoxing Xu*†, I. Scott Ramsey*†, Suhas A. Kotecha*†,
Magdalene M. Moran‡, Jayhong A. Chong*, Deborah Lawson§, Pei Ge§, Jeremiah Lilly§, Inmaculada Silos-Santiago§, Yu Xie§,
Peter S. DiStefano§k, Rory Curtis§k & David E. Clapham*

* Howard Hughes Medical Institute, Children’s Hospital, Harvard Medical School, Enders 1309, 320 Longwood Avenue, Boston, Massachusetts 02115, USA ‡ Program in Neuroscience, Harvard Medical School, Goldenson Building B2-228, 220 Longwood Avenue, Boston, Massachusetts 02115, USA
§ Millennium Pharmaceuticals Inc., 75 Sidney Street, Cambridge, Massachusetts 02139, USA
† These authors contributed equally to this work

Transient receptor potential (TRP) proteins are cation-selective channels that function in processes as diverse as sensation and vasoregulation. Mammalian TRP channels that are gated by heat and capsaicin (>43 8C; TRPV1 (ref. 1)), noxious heat (>52 8C; TRPV2 (ref. 2)), and cooling (< 22 8C; TRPM8 (refs 3, 4)) have been cloned; however, little is known about the molecular determinants of temperature sensing in the range between ,22 8C and 40 8C. Here we have identified a member of the vanilloid channel family, human TRPV3 (hTRPV3) that is expressed in skin, tongue, dorsal root ganglion, trigeminal ganglion, spinal cord and brain. Increasing temperature from 228C to 408C in mammalian cells transfected with hTRPV3 elevated intracellular calcium by activating a nonselective cat- ionic conductance. As in published recordings from sensory neurons, the current was steeply dependent on temperature, sensitized with repeated heating, and displayed a marked hyster- esis on heating and cooling5–10. On the basis of these properties, we propose that hTRPV3 is thermosensitive in the physiological range of temperatures between TRPM8 and TRPV1.

Journal Club 2014-03-28 Read More »

Journal Club 2014-03-22

Mustard oils and cannabinoids excite sensory nerve fibres through the TRP channel ANKTM1

nature02282

Sven-Eric Jordt1, Diana M. Bautista1, Huai-hu Chuang1,
David D. McKemy1, Peter M. Zygmunt3, Edward D. Ho ̈ gesta ̈ tt3, Ian D. Meng2* & David Julius1

1Department of Cellular and Molecular Pharmacology and 2Department of Neurology, University of California, San Francisco, California 94143-2140, USA 3Department of Clinical Pharmacology, Institute of Laboratory Medicine,
Lund University Hospital, SE-221 85 Lund, Sweden

* Present address: Department of Physiology, College of Osteopathic Medicine, University of New England, 11 Hills Beach Road, Biddeford, Maine 04005, USA ………………………………………………………………………………………………………………………………………………………..

Wasabi, horseradish and mustard owe their pungency to iso- thiocyanate compounds. Topical application of mustard oil (allyl isothiocyanate) to the skin activates underlying sensory nerve endings, thereby producing pain, inflammation and robust

1,2 hypersensitivity to thermal and mechanical stimuli . Despite

their widespread use in both the kitchen and the laboratory, the molecular mechanism through which isothiocyanates mediate their effects remains unknown. Here we show that mustard oil depolarizes a subpopulation of primary sensory neurons that are also activated by capsaicin, the pungent ingredient in chilli peppers, and by D9-tetrahydrocannabinol (THC), the psycho- active component of marijuana. Both allyl isothiocyanate and THC mediate their excitatory effects by activating ANKTM1, a member of the transient receptor potential (TRP) ion channel family recently implicated in the detection of noxious cold3,4. These findings identify a cellular and molecular target for the pungent action of mustard oils and support an emerging role for TRP channels as ionotropic cannabinoid receptors5–8.

Journal Club 2014-03-22 Read More »

Journal Club 2014.03.05

Enhanced excitability of MRGPRA3- and MRGPRD-positive nociceptors in a model ofinflammatory itch and pain

Lintao Qu,1,* Ni Fan,1,2,* Chao Ma,1,3 Tao Wang,1,3 Liang Han,4 Kai Fu,1 Yingdi Wang,5 Steven G. Shimada,1 Xinzhong Dong4 and Robert H. LaMotte1

  1. 1  Department of Anaesthesiology, Yale University School of Medicine, New Haven, CT, 06520, USA
  2. 2  Guangzhou Brain Hospital, the Affiliated Hospital of Guangzhou Medical University, Guangzhou, Guangdong, China 510370
  3. 3 InstituteofBasicMedicalSciences,Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, Department of Anatomy, Histology and Embryology, Beijing, China
  4. 4  Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA
  5. 5  Section of Cardiovascular Medicine, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06511, USA

*These authors contributed equally to this work.

Correspondence to: Robert H. LaMotte, Ph.D.,
Department of Anaesthesiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA
E-mail: robert.lamotte@yale.edu

Brain-2014-Qu-brain_awu007

Itch is a common symptom of diseases of the skin but can also accompany diseases of other tissues including the nervous system. Acute itch from chemicals experimentally applied to the skin is initiated and maintained by action potential activity in a subset of nociceptive neurons. But whether these pruriceptive neurons are active or might become intrinsically more excitable under the pathological conditions that produce persistent itch and nociceptive sensations in humans is largely unexplored. Recently, two distinct types of cutaneous nociceptive dorsal root ganglion neurons were identified as responding to pruritic chemicals and playing a role in itch sensation. One expressed the mas-related G-coupled protein receptor MRGPRA3 and the other MRGPRD (MRGPRA3+ and MRGPRD+ neurons, respectively). Here we tested whether these two distinct pruriceptive nociceptors exhibited an enhanced excitability after the development of contact hypersensitivity, an animal model of allergic contact dermatitis, a common pruritic disorder in humans. The characteristics of increased excitability of pruriceptive neurons during this disorder may also pertain to the same types of neurons active in other pruritic diseases or pathologies that affect the nervous system and other tissues or organs. We found that challenging the skin of the calf of the hind paw or the cheek of previously sensitized mice with the hapten, squaric acid dibutyl ester, produced symptoms of contact hypersensitivity including an increase in skin thickness and site-directed spontaneous pain-like (licking or wiping) and itch-like (biting or scratching) behaviours. Ablation of MRGPRA3+ neurons led to a significant reduction in spontaneous scratching of the hapten-challenged nape of the neck of previously sensitized mice. In vivo, electrophysiological recordings revealed that MRGPRA3+ and MRGPRD+ neurons innervating the hapten-challenged skin exhibited a greater incidence of spontaneous activity and/or abnor- mal after-discharges in response to mechanical and heat stimuli applied to their receptive fields compared with neurons from the vehicle-treated control animals. Whole-cell recordings in vitro showed that both MRGPRA3+ and MRGPRD+ neurons from hapten-challenged mice displayed a significantly more depolarized resting membrane potential, decreased rheobase, and greater number of action potentials at twice rheobase compared with neurons from vehicle controls. These signs of neuronal hyper- excitability were associated with a significant increase in the peak amplitude of tetrodotoxin-sensitive and resistant sodium currents. Thus, the hyperexcitability of MRGPRA3+ and MRGPRD+ neurons, brought about in part by enhanced sodium currents, may contribute to the spontaneous itch- and pain-related behaviours accompanying contact hypersensitivity and/or other inflammatory diseases in humans.

Keywords: itch; pain; MRGPRA3; MRGPRD; allergic contact dermatitis
Abbreviations: CHS = contact hypersensitivity; GFP = green fluorescent protein; SADBE = squaric acid dibutyl ester;

TTX = tetrodotoxin

Journal Club 2014.03.05 Read More »

Journal club 2014.02.13.

Cortical GluK1 kainate receptors modulate scratching in adult mice

Journal of Neurochemistry | 2013 | 126 | 636-650

Kaori Yamada and Min Zhuo

jnc12351

Abstract

Recent investigations into the mechanisms mediating itch transmission have focused on spinal mechanisms, whereas few studies have investigated the role of the cerebral cortex in itch-related behaviors. Human imaging studies show that several cortical regions are active in correspondence with itch, including the anterior cingulate cortex (ACC). We present here evidence of cortical modulation of pruritogen-induced scratch- ing behavior. We combine pharmacological, genetic, and electrophysiological approaches to show that cortical GluK1- containing kainate (KA) receptors are involved in scratching

induced by histamine and non-histamine-dependent itching stimuli. We further show that scratching corresponds with enhanced excitatory transmission in the ACC through KA receptor modulation of inhibitory circuitry. In addition, we found that inhibiting GluK1-containing KA receptors in the ACC also reduced behavioral nociceptive responses induced by forma- lin. Our results reveal a new role of the cortex in pruritogen- induced scratching.

Keywords: cortex, glutamatergic transmission, inhibitory trans- mission, itch, kainate receptors, scratching.
J. Neurochem. (2013) 126, 636–650.

Journal club 2014.02.13. Read More »

Journal club 2014.01.29.

Distinct Expression of Mas1-Related G-Protein-Coupled Receptor B4 in Dorsal Root and Trigeminal Ganglia—Implications for Altered Behaviors in Acid-Sensing Ion Channel 3-Deficient Mice

Ya-Han Huang & Chin-Yu Chang & Chih-Cheng Chen & Chih-Dong Yang & Wei-Hsin Sun

Received: 26 February 2013 / Accepted: 8 July 2013 / Published online: 31 July 2013 # Springer Science+Business Media New York 2013

art%3A10.1007%2Fs12031-013-0070-0

Abstract

Mas1-related G-protein-coupled receptors (Mrgprs), comprising more than 50 distinct members, are specifically expressed in primary sensory neurons. Reflecting the diversity and specificity of stimuli they detect, Mrgprs are involved in pain, touch, and itch-related behaviors. Sensory–neuron-specif- ic acid-sensing ion channel 3 (ASIC3) is essential for touch and inflammatory pain, but mice lacking ASIC3 have complex behavioral alterations in various modalities of pain and touch. To understand whether Mrgprs are involved in complex behav- ioral alterations found in ASIC3-deficient mice, we examined Mrgpr gene expression in ASIC3−/− mice. Only MrgprB4 expression has shown significant change. MrgprB4 expression was increased in ASIC3−/− dorsal root ganglia (DRG) but decreased in ASIC3−/− trigeminal ganglia. The distinct alter- ations in DRG and trigeminal ganglia imply that MrgprB4 could have multiple functions. Given that MrgprB4 is expressed in neurons that may detect gentle touch and that ASIC3−/− mice have altered sensitivity of mechanoreceptors for light touch, the expression change of MrgprB4 is more likely related to the altered touch behaviors of ASIC3−/− mice.

Keywords

Dorsal root ganglion . Trigeminal ganglion . Mas1-related G-protein-coupled receptor B4 . ASIC3 . Nociceptors

Journal club 2014.01.29. Read More »

Journal Club 2014.01.16

Roles of glutamate, substance P, and gastrin-releasing peptide as spinal neurotransmitters of histaminergic and nonhistaminergic itch

Tasuku Akiyama a, Mitsutoshi Tominaga b, Kenji Takamori b, Mirela Iodi Carstens a, E. Carstens a,⇑ a Department of Neurobiology, Physiology & Behavior, University of California, Davis, CA, USA

b Institute for Environmental and Gender Specific Medicine, Juntendo University Graduate School of Medicine, Urayasu, Chiba 279-0021, Japan

1-s2.0-S0304395913005083-main

abstract

We investigated roles for substance P (SP), gastrin-releasing peptide (GRP), and glutamate in the spinal neurotransmission of histamine-dependent and -independent itch. In anesthetized mice, responses of single superficial dorsal horn neurons to intradermal (i.d.) injection of chloroquine were partially reduced by spinal application of the a-amino-3-hydroxy-5-methyl-4-isoxazole proprionate acid (AMPA)/kainate antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Co-application of CNQX plus a neurokinin-1 (NK-1) antagonist produced stronger inhibition, while co-application of CNQX, NK-1, and GRP receptor (GRPR) antagonists completely inhibited firing. Nociceptive-specific and wide dynamic range-type neu- rons exhibited differential suppression by CNQX plus either the GRPR or NK-1 antagonist, respectively. Neuronal responses elicited by i.d. histamine were abolished by CNQX alone. In behavioral studies, indi- vidual intrathecal administration of a GRPR, NK-1, or AMPA antagonist each significantly attenuated chlo- roquine-evoked scratching behavior. Co-administration of the NK-1 and AMPA antagonists was more effective, and administration of all 3 antagonists abolished scratching. Intrathecal CNQX alone prevented histamine-evoked scratching behavior. We additionally employed a double-label strategy to investigate molecular markers of pruritogen-sensitive dorsal root ganglion (DRG) cells. DRG cells responsive to his- tamine and/or chloroquine, identified by calcium imaging, were then processed for co-expression of SP, GRP, or vesicular glutamate transporter type 2 (VGLUT2) immunofluorescence. Subpopulations of chloro- quine- and/or histamine-sensitive DRG cells were immunopositive for SP and/or GRP, with >80% immu- nopositive for VGLUT2. These results indicate that SP, GRP, and glutamate each partially contribute to histamine-independent itch. Histamine-evoked itch is mediated primarily by glutamate, with GRP play- ing a lesser role. Co-application of NK-1, GRP, and AMPA receptor antagonists may prove beneficial in treating chronic itch.

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Journal Club 2013/12/26

Modulation of Transient Receptor Vanilloid 1 Activity by Transient Receptor Potential Ankyrin 1
Viola Spahn, Christoph Stein and Christian Zöllner

Transient Receptor Potential Vanilloid 1 (TRPV1) is a nonselective ligand-gated cation channel responding to noxious heat, protons, and chemicals like capsaicin. TRPV1 is expressed in sensory neurons and plays a critical role in pain associated with tissue injury, inflammation or nerve lesions. Transient Receptor Potential Ankyrin 1 (TRPA1) is co- expressed with TRPV1. It is activated by compounds that cause a burning sensation (e.g. mustard oil) and, indirectly, by components of the inflammatory milieu eliciting nociceptor excitation and pain hypersensitivity. Previous studies indicate an interaction of TRPV1 and TRP A1 signaling pathways. Here we sought to examine the molecular mechanisms underlying such interactions in nociceptive neurons. We first excluded physical interactions of both channels using radioligand binding studies. By microfluorimetry, electrophysiological experiments, cAMP measurements, and site-directed mutagenesis we found a sensitization of TRPV1 after TRPA1 stimulation with mustard oil in a calcium- and cAMP/PKA-dependent manner. TRPA1 stimulation enhanced TRPV1 phosphorylation via the putative PKA phosphorylation site serine 116. We also detected calcium-sensitive increased TRPV1 activity after TRPA1 activation in dorsal root ganglion (DRG) neurons. The inhibition of TRPA1 by HC-030031 after its initial stimulation and the calcium-insensitive TRPA1 mutant D477A still showed increased capsaicin-induced TRPV1 activity excluding an additive TRPA1 current after TRPV1 stimulation. Our study shows sensitization of TRPV1 via activation of TRPA1, which involves adenylyl cyclase, increased cAMP, subsequent translocation and activation of PKA, and phosphorylation of TRPV1 at PKA phosphorylation residues. This suggests that cross-sensitization of TRP channels contributes to enhanced pain sensitivity in inflamed tissues.

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