Journal Club 18.02.09.

Pharmacological evidence of involvement of nitric oxide pathway in anti-pruritic effects of sumatriptan in chloroquine-induced scratching in mice

Pharmacological evidence of involvement of nitric oxide pathway in anti-pruritic effects of sumatriptan in chloroquine-induced scratching in mice

Nazgol-Sadat Haddadia,b, Sattar Ostadhadia,b,c, Saeed Shakibaa,b, Khashayar Afsharia,b, Nastaran Rahimia,b, Arash Foroutana,b, Ahmad-Reza Dehpoura,b* aExperimental Medicine Research Center, Tehran University of Medical Sciences, Poorsina St., Enghelab Ave., Tehran, Iran bDepartment of Pharmacology, School of Medicine, Tehran University of Medical Sciences, Poorsina St., Enghelab Ave., Tehran, Iran cBrain and Spinal Cord Injury Research Center, Neuroscience Institute, Tehran University of Medical Sciences, Chamran highway, Bagherkhan St., Imam Khomeini Hospital, Tehran, Iran

Chloroquine (CQ) induces histamine-independent itch in human and mice. We recently reported the role of intradermal nitric oxide (NO)/cyclic guanosine monophosphate pathway in CQ-evoked scratching in mice. Chloroquine stimulates neuronal nitric oxide synthase (nNOS) activity to over-producing NO in the skin. Sumatriptan, a 5-hydroxytryptamine 1b/1d receptors (5-HTR1b/1d) agonist, is involved in pain and used to treat migraine and cluster headaches. According to previous studies, sumatriptan inhibits NOS activity. Thus, we aimed to investigate the effect of sumatriptan on CQ-induced scratching. We used the rostral back model of itch. Chloroquine was injected intradermally into the rostral back of NMRI mice, and the scratching behavior was evaluated by measuring the number of bouts over 30 min. We evaluated the effect of sumatriptan and combination of sumatriptan and a non-selective NO synthase inhibitor, L-N-nitro arginine methyl ester (L-NAME), on the scratching behavior. Additionally, the changes of skin, hip- pocampus, and cortical nitrite level after different treatments were studied. Intraperitoneal and intradermal sumatriptan attenuates CQ-induced itch which reversed by GR-127935, the selective 5-HTR1b and 5-HTR1d antagonist. Co- administration of subeffective doses of sumatriptan and L-NAME significantly decreases the scratching behavior. Intradermal injection of CQ significantly increases the intradermal nitrite levels while it does not have any significant effects on hippocampal or cortical nitrite concentrations. Likewise, the effective doses of intraperitoneal and intradermal sumatriptan significantly reduce intradermal nitrite levels. We concluded that sumatriptan suppresses CQ-induced itch most likely by activating 5-HT1b/1d receptors. This effect probably mediates through NO pathway.

Journal Club 18.02.09. Read More »

2018.2.2

TRPV3 Channel in Keratinocytes in Scars with Post-Burn Pruritus.

Park CW, et al. Int J Mol Sci. 2017.

Abstract

Post-burn pruritus is a common and distressing sequela of burn scars. Empirical antipruritic treatments usually fail to have a satisfactory outcome because of their limited selectivity and possible side effects. Therefore, novel drug targets need to be identified. Here, we aimed to investigate the possible role of protease-activated receptor 2 (PAR2) and transient receptor potential vanniloid 3 (TRPV3), along with the relation of TRPV3 to thymic stromal lymphopoietin (TSLP). Specimens from normal (unscarred) or burn-scarred (with or without pruritus) tissue were obtained from burn patients for this study. In each sample, the keratinocytes were isolated and cultured, and the intracellular Ca2+ level at the time of stimulation of each factor was quantified and the interaction was screened. PAR2 function was reduced by antagonism of TRPV3. Inhibiting protein kinase A (PKA) and protein kinase C (PKC) reduced TRPV3 function. TSLP mRNA and protein, and TSLPR protein expressions, increased in scars with post-burn pruritus, compared to scars without it or to normal tissues. In addition, TRPV1 or TRPV3 activation induced increased TSLP expression. Conclusively, TRPV3 may contribute to pruritus in burn scars through TSLP, and can be considered a potential therapeutic target for post-burn pruritus.

20180202 YSH

2018.2.2 Read More »

2018.01.26

P2Y1 Receptor Activation of the TRPV4 Ion Channel Enhances Purinergic Signaling in Satellite Glial Cells.

Rajasekhar P1, Poole DP2, Liedtke W3, Bunnett NW4, Veldhuis NA5.

Abstract

Transient receptor potential (TRP) ion channels of peripheral sensory pathways are important mediators of pain, itch, and neurogenic inflammation. They are expressed by primary sensory neurons and by glial cells in the central nervous system, but their expression and function in satellite glial cells (SGCs) of sensory ganglia have not been explored. SGCs tightly ensheath neurons of sensory ganglia and can regulate neuronal excitability in pain and inflammatory states. Using a modified dissociation protocol, we isolated neurons with attached SGCs from dorsal root ganglia of mice. SGCs, which were identified by expression of immunoreactive Kir4.1 and glutamine synthetase, were closely associated with neurons, identified using the pan-neuronal marker NeuN. A subpopulation of SGCs expressed immunoreactive TRP vanilloid 4 (TRPV4) and responded to the TRPV4-selective agonist GSK1016790A by an influx of Ca(2+) ions. SGCs did not express functional TRPV1, TRPV3, or TRP ankyrin 1 channels. Responses to GSK1016790A were abolished by the TRPV4 antagonist HC067047 and were absent in SGCs from Trpv4(-/-) mice. The P2Y1-selective agonist 2-methylthio-ADP increased [Ca(2+)]i in SGCs, and responses were prevented by the P2Y1-selective antagonist MRS2500. P2Y1 receptor-mediated responses were enhanced in TRPV4-expressing SGCs and HEK293 cells, suggesting that P2Y1 couples to and activates TRPV4. PKC inhibitors prevented P2Y1 receptor activation of TRPV4. Our results provide the first evidence for expression of TRPV4 in SGCs and demonstrate that TRPV4 is a purinergic receptor-operated channel in SGCs of sensory ganglia.

18:01:26

2018.01.26 Read More »

2018.01.19

Distinct roles of NMB and GRP in itch transmission.

Wan L1,2,3, Jin H1,2,4, Liu XY1,2, Jeffry J1,2, Barry DM1,2, Shen KF1,2,5, Peng JH1,2, Liu XT1,2,6, Jin JH1,2,7, Sun Y1,2,8, Kim R1, Meng QT1,2,9, Mo P1,2,10, Yin J1,2, Tao A6, Bardoni R11, Chen ZF12,13,14,15.

Abstract

A key question in our understanding of itch coding mechanisms is whether itch is relayed by dedicated molecular and neuronal pathways. Previous studies suggested that gastrin-releasing peptide (GRP) is an itch-specific neurotransmitter. Neuromedin B (NMB) is a mammalian member of the bombesin family of peptides closely related to GRP, but its role in itch is unclear. Here, we show that itch deficits in micelacking NMB or GRP are non-redundant and Nmb/Grp double KO (DKO) mice displayed additive deficits. Furthermore, both Nmb/Grp and Nmbr/Grpr DKO mice responded normally to a wide array of noxious stimuli. Ablation of NMBR neurons partially attenuated peripherally induced itch without compromising nociceptive processing. Importantly, electrophysiological studies suggested that GRPR neurons receive glutamatergic input from NMBR neurons. Thus, we propose that NMB and GRP may transmit discrete itch information and NMBR neurons are an integral part of neural circuits for itch in the spinal cord.

Distinct roles of NMB and GRP in itch transmission.

Supplementary information

2018.01.19 Read More »

Journal Club 18.01.12.

Transient receptor potential vanilloid 4–expressing macrophages and keratinocytes contribute differentially to allergic and nonallergic chronic itch

TRP4-expressing macrophages and keratinocytes contribute differentially to allergic and nonallergic chronic itch

Jialie Luo, PhD,a* Jing Feng, PhD,a* Guang Yu, PhD,a,b Pu Yang, PhD,a Madison R. Mack, BA,a Junhui Du, PhD,c Weihua Yu, PhD,d Aihua Qian, PhD,e Yujin Zhang, PhD,f Shenbin Liu, PhD,a Shijin Yin, PhD,g Amy Xu, BS,a Jizhong Cheng, PhD,h Qingyun Liu, PhD,i Roger G. O’Neil, PhD,j Yang Xia, PhD,f Liang Ma, PhD,k
Susan M. Carlton, PhD,c Brian S. Kim, MD,a,k Kenneth Renner, PhD,l Qin Liu, PhD,a and Hongzhen Hu, PhDa
St Louis, Mo; Nanjing, Chongqing, Shanghai, and Wuhan, China; Galveston and Houston, Tex; and Vermillion, SD

Background: Chronic itch is a highly debilitating symptom that underlies many medical disorders with no universally effective treatments. Although unique neuronal signaling cascades in the sensory ganglia and spinal cord have been shown to critically promote the pathogenesis of chronic itch, the role of skin- associated cells remains poorly understood.
Objective: We sought to examine the cutaneous mechanisms underlying transient receptor potential vanilloid 4 (TRPV4)–mediated allergic and nonallergic chronic itch. Methods: Expression of TRPV4 in chronic itch and healthy control skin preparations was examined by using real-time RT-PCR. Trpv4eGFP mice were used to study the expression and function of TRPV4 in the skin by means of immunofluorescence staining, flow cytometry, calcium imaging, and patch-clamp recordings. Genetic and pharmacologic approaches were used to examine the role and underlying mechanisms of TRPV4 in mouse models of dry skin–associated chronic itch and spontaneous scratching associated with squaric acid dibutylester–induced allergic contact dermatitis.
Results: TRPV4 is selectively expressed by dermal macrophages and epidermal keratinocytes in mice. Lineage-specific deletion of TRPV4 in macrophages and keratinocytes reduces allergic and nonallergic chronic itch in mice, respectively. Importantly, TRPV4 expression is significantly increased in skin biopsy specimens from patients with chronic idiopathic pruritus in comparison with skin from healthy control subjects. Moreover, TRPV4-dependent chronic itch requires 5-hydroxytryptamine (5-HT) signaling secondary to activation of distinct 5-HT receptors in both patients with allergic and those with nonallergic chronic itch conditions.
Conclusion: Our study reveals previously unrecognized mechanisms by which TRPV4-expressing epithelial and immune cells in the skin critically and dynamically mediate chronic itch and unravels novel targets for therapeutics in the setting of chronic itch. (J Allergy Clin Immunol 2017;nnn:nnn-nnn.)
Key words: Transient receptor potential vanilloid 4, chronic itch, macrophage, keratinocyte

Journal Club 18.01.12. Read More »

2018.01.05

UVB radiation generates sunburn pain and affects skin by activating epidermal TRPV4 ion channels and triggering endothelin-1 signaling.

Moore C1, Cevikbas F, Pasolli HA, Chen Y, Kong W, Kempkes C, Parekh P, Lee SH, Kontchou NA, Yeh I, Jokerst NM, Fuchs E, Steinhoff M, Liedtke WB.

Abstract

At our body surface, the epidermis absorbs UV radiation. UV overexposure leads to sunburn with tissue injury and pain. To understand how, we focus on TRPV4, a nonselective cation channel highly expressed in epithelial skin cells and known to function in sensory transduction, a property shared with other transient receptor potential channels. We show that following UVB exposure mice with induced Trpv4 deletions, specifically in keratinocytes, are less sensitive to noxious thermal and mechanical stimuli than control animals. Exploring the mechanism, we find that epidermal TRPV4 orchestrates UVB-evoked skin tissue damage and increased expression of the proalgesic/algogenic mediator endothelin-1. In culture, UVB causes a direct, TRPV4-dependent Ca(2+) response in keratinocytes. In mice, topical treatment with a TRPV4-selective inhibitor decreases UVB-evoked pain behavior, epidermal tissue damage, and endothelin-1 expression. In humans, sunburn enhances epidermal expression of TRPV4 and endothelin-1, underscoring the potential of keratinocyte-derived TRPV4 as a therapeutic target for UVB-induced sunburn, in particular pain.

1월 UVB radiation generates sunburn pain and affects skin by activating epidermal TRPV4 ion channels and triggering endothelin-1 signaling

supplimentary

2018.01.05 Read More »

2017.12.29

Sensory TRP channels contribute differentially to skin inflammation and persistent itch.

Feng J1, Yang P1, Mack MR2, Dryn D1,3, Luo J1, Gong X1,4, Liu S1, Oetjen LK2, Zholos AV3, Mei Z4, Yin S4, Kim BS2, Hu H5.

Abstract

Although both persistent itch and inflammation are commonly associated with allergic contact dermatitis (ACD), it is not known if they are mediated by shared or distinct signaling pathways. Here we show that both TRPA1 and TRPV1 channels are required for generating spontaneous scratching in a mouse model of ACD induced by squaric acid dibutylester (SADBE), a small molecule hapten, through directly promoting the excitability of pruriceptors. TRPV1 but not TRPA1 channels protect the skin inflammation, as genetic ablation of TRPV1 function or pharmacological ablation of TRPV1-positive sensory nerves promotes cutaneous inflammation in the SADBE-induced ACD. Our results demonstrate that persistent itch and inflammation are mediated by distinct cellular and molecular mechanisms in a mouse model of ACD. Identification of distinct roles of TRPA1 and TRPV1 in regulating itch and inflammation may provide new insights into the pathophysiology and treatment of chronic itch and inflammation in ACD patients.

Sensory TRP channels contribute differentially to skin inflammation and persistent itch.

Supplementary information

2017.12.29 Read More »

Journal Club 17.12.22.

TRPC3 Is Dispensable for β-Alanine Triggered Acute Itch
Peter Dong1, Changxiong Guo2, Shengxiang Huang2,3, Minghong Ma1, Qin Liu2 & Wenqin Luo1

TRPC3 Is Dispensable for β-Alanine Triggered Acute Itch

The detection of pruritic (itchy) stimuli is mediated by a variety of receptors and channels expressed
by primary sensory neurons. The G protein-coupled receptor (GPCR) MRGPRD is selectively expressed by a subset of mouse non-peptidergic nociceptors and functions as the molecular receptor for the itch-inducing chemical β-alanine. However, the channels responsible for generating electrical signals downstream of MRGPRD remain unclear. Here, we found that a member of the canonical TRP channel family, TRPC3, is highly expressed in MRGPRD+ non-peptidergic nociceptors, raising the possibility of whether TRPC3 functions as a downstream channel in the MRGPRD signaling pathway. We tested TrpC3 null mice for β-alanine induced itch, and found that these mice exhibit normal responses to β-alanine. At the cellular level, calcium in ux triggered by β-alanine is also unchanged in cultured DRG neurons from TrpC3 null mice compared to wild type. Together, our results demonstrate that mouse TrpC3 is dispensable for β-alanine-induced acute itch.

Journal Club 17.12.22. Read More »

2017.12.15

Transient Receptor Potential Vanilloid 4 Ion Channel Functions as a Pruriceptor in Epidermal Keratinocytes to Evoke Histaminergic Itch*

Abstract

TRPV4 ion channels function in epidermal keratinocytes and in innervating sensory neurons; however, the contribution of the channel in either cell to neurosensory function remains to be elucidated. We recently reported TRPV4 as a critical component of the keratinocyte machinery that responds to ultraviolet B (UVB) and functions critically to convert the keratinocyte into a pain-generator cell after excess UVB exposure. One key mechanism in keratinocytes was increased expression and secretion of endothelin-1, which is also a known pruritogen. Here we address the question of whether TRPV4 in skin keratinocytes functions in itch, as a particular form of “forefront” signaling in non-neural cells. Our results support this novel concept based on attenuated scratching behavior in response to histaminergic (histamine, compound 48/80, endothelin-1), not non-histaminergic (chloroquine) pruritogens in Trpv4 keratinocyte-specific and inducible knock-out mice. We demonstrate that keratinocytes rely on TRPV4 for calcium influx in response to histaminergic pruritogens. TRPV4 activation in keratinocytes evokes phosphorylation of mitogen-activated protein kinase, ERK, for histaminergic pruritogens. This finding is relevant because we observed robust anti-pruritic effects with topical applications of selective inhibitors for TRPV4 and also for MEK, the kinase upstream of ERK, suggesting that calcium influx via TRPV4 in keratinocytes leads to ERK-phosphorylation, which in turn rapidly converts the keratinocyte into an organismal itch-generator cell. In support of this concept we found that scratching behavior, evoked by direct intradermal activation of TRPV4, was critically dependent on TRPV4 expression in keratinocytes. Thus, TRPV4 functions as a pruriceptor-TRP in skin keratinocytes in histaminergic itch, a novel basic concept with translational-medical relevance.

Transient Receptor Potential Vanilloid 4 Ion Channel Functions as a Pruriceptor in Epidermal Keratinocytes to Evoke Histaminergic Itch

2017.12.15 Read More »

2017.12.08

Targeting human Mas-related G protein-coupled receptor X1 to inhibit persistent pain.

Li Z1, Tseng PY1, Tiwari V2, Xu Q1, He SQ2, Wang Y1, Zheng Q1, Han L1, Wu Z3,4, Blobaum AL5, Cui Y6, Tiwari V2, Sun S1, Cheng Y1, Huang-Lionnet JH2, Geng Y1, Xiao B6, Peng J3,4,7, Hopkins C5, Raja SN2, Guan Y8, Dong X9,10.

Abstract

Human Mas-related G protein-coupled receptor X1 (MRGPRX1) is a promising target for pain inhibition, mainly because of its restricted expression in nociceptors within the peripheral nervous system. However, constrained by species differences across Mrgprs, drug candidates that activate MRGPRX1 do not activate rodent receptors, leaving no responsive animal model to test the effect on pain in vivo. Here, we generated a transgenic mouse line in which we replaced mouse Mrgprs with human MrgprX1 This humanized mouse allowed us to characterize an agonist [bovine adrenal medulla 8-22 (BAM8-22)] and a positive allosteric modulator (PAM), ML382, of MRGPRX1. Cellular studies suggested that ML382 enhances the ability of BAM8-22 to inhibit high-voltage-activated Ca2+ channels and attenuate spinal nociceptive transmission. Importantly, both BAM8-22 and ML382 effectively attenuated evoked, persistent, and spontaneous pain without causing obvious side effects. Notably, ML382 by itself attenuated both evoked pain hypersensitivity and spontaneous pain in MrgprX1 mice after nerve injury without acquiring coadministration of an exogenous agonist. Our findings suggest that humanized MrgprX1 mice provide a promising preclinical model and that activating MRGPRX1 is an effective way to treat persistent pain.

KEYWORDS: DRG neurons; GPCR; MrgprX1; pain; positive allosteric modulator

Targeting human Mas-related G protein coupled receptor X1 to inhibit persistent pain.

2017.12.08 Read More »

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