Itch Behavior Research Articles

Innocuous warming enhances peripheral serotonergic itch signaling and evokes enhanced responses in serotonin-responsive dorsal horn neurons in the mouse.

Skin warming often exacerbates itch in patients with itchy dermatitis. We demonstrate that warming the skin enhanced serotonin-evoked, but not histamine-evoked, itch behavior and responses of sensory dorsal root ganglion cells. Moreover, serotonin, but not histamine, enhanced responses of superficial dorsal horn neurons to innocuous warming. The results suggest that skin warming selectively enhances the responses of serotonin-sensitive pruriceptors, leading to increased firing of serotonin-sensitive dorsal horn neurons that signal nonhistaminergic itch.

Mast cell proteases protect against histaminergic itch and attenuate tissue injury pain responses

Abstract Aims Mast cell proteases comprise up to 25% of the total protein content within mast cells and have various functions that are still poorly understood but can be either protective or pro-inflammatory.Three ofthose proteases in mice have been identified as the closest functional homologues to human mast cell proteases, and in this currently ongoing study we are investigating if they play a role in itch and tissue injury pain, and if they do, what the possible mechanism behind that could be. Methods Knock-out mice were generated, lacking either one or multiple proteases. To evaluate itch, the animals (n = 8–10) were injected intradermally with histaminergic (histamine, compound 48/80) and non-histaminergic (protease activated receptor 2 (PAR2) agonist, chloroquine) pruritic substances and their itch behavior over 1 h scored. Formalin injection (n = 20) in the hind paw was used as a model for tissue injury pain, where the mice were monitored for pain behavior for 1 h following the injection. Age and gender matched C57BL/6 mice were used as controls. Results Mice lacking three of the proteases had twice as many scratching episodes in response to histamine (P =0.016) and compound 48/80 (P =0.034) than controls. The non-histaminergic substances chloroqine and PAR2 are more potent pruritogens but difference was not seen in itch levels between genotypes. Furthermore, the protease-deficient mice exhibited a more pronounced pain response during the inflammatory stage (10–60 min after formalin injection, P = 0.025) of the tissue injury test. Conclusions The mouse mast cell proteases have a protective role against histaminergic itch and in acute inflammation following a tissue injury. The exact mechanism remains unclear at present as the proteases cleave a variety of different pro-inflammatory substrates and can act in complimentary fashion by cleaving the same substrates but at different sites.

Identification of a Neuronal Receptor Controlling Anaphylaxis.

Allergic reactions can in severe cases induce a state of circulatory shock referred to as anaphylaxis. Histamine, the primary mediator of this condition, is released from immune cells, and, therefore, anaphylaxis has so far been considered an immune system disorder. However, we here show that the glutamatergic receptor mGluR7, expressed on a subpopulation of both peripheral and spinal cord neurons, controls histamine-induced communication through calcium-dependent autoinhibition with implications for anaphylaxis. Genetic ablation of mGluR7, and thus altered regulation of histamine-sensing neurons, caused an anaphylaxis-like state in mGluR7(-/-) mice, which could be reversed by antagonizing signaling between neurons and mast cells but not by antagonizing a central itch pathway. Our findings demonstrate the vital role of nervous system control by mGluR7 in anaphylaxis and open up possibilities for preventive strategies for this life-threatening condition.

TRPV1 and PLC Participate in Histamine H4 Receptor-Induced Itch

Histamine H4 receptor has been confirmed to play a role in evoking peripheral pruritus. However, the ionic and intracellular signaling mechanism of activation of H4 receptor on the dorsal root ganglion (DRG) neurons is still unknown. By using cell culture and calcium imaging, we studied the underlying mechanism of activation of H4 receptor on the DRG neuron. Immepip dihydrobromide (immepip)—a histamine H4 receptor special agonist under cutaneous injection—obviously induced itch behavior of mice. Immepip-induced scratching behavior could be blocked by TRPV1 antagonist AMG9810 and PLC pathway inhibitor U73122. Application of immepip (8.3–50 μM) could also induce a dose-dependent increase in intracellular Ca2+ ([Ca2+]i) of DRG neurons. We found that 77.8% of the immepip-sensitized DRG neurons respond to the TRPV1 selective agonist capsaicin. U73122 could inhibit immepip-induced Ca2+ responses. In addition, immepip-induced [Ca2+]i increase could be blocked by ruthenium red, capsazepine, and AMG9810; however it could not be blocked by TRPA1 antagonist HC-030031. These results indicate that TRPV1 but not TRPA1 is the important ion channel to induce the DRG neurons' responses in the downstream signaling pathway of histamine H4 receptor and suggest that TRPV1 may be involved in the mechanism of histamine-induced itch response by H4 receptor activation.

Gate control of mechanical itch by a subpopulation of spinal cord interneurons

Light mechanical stimulation of hairy skin can induce a form of itch known as mechanical itch. This itch sensation is normally suppressed by inputs from mechanoreceptors; however, in many forms of chronic itch, including alloknesis, this gating mechanism is lost. Here we demonstrate that a population of spinal inhibitory interneurons that are defined by the expression of neuropeptide Y::Cre (NPY::Cre) act to gate mechanical itch. Mice in which dorsal NPY::Cre-derived neurons are selectively ablated or silenced develop mechanical itch without an increase in sensitivity to chemical itch or pain. This chronic itch state is histamine-independent and is transmitted independently of neurons that express the gastrin-releasing peptide receptor. Thus, our studies reveal a dedicated spinal cord inhibitory pathway that gates the transmission of mechanical itch.

A circuit controlling mechanical itch

Chronic Itch Considerable progress has been made in understanding and treating chemically induced itch. However, little is known about the mechanisms underlying mechanically evoked itch. Bourane et al. produced a model of mechanical itch by reducing the number of neuropeptide Y–expressing inhibitory spinal interneurons. This led to a selective increase in mechanically evoked itch-like behavior in mice. In contrast, chemically evoked itch or pain behavior remained unaffected. Science , this issue p. [550][1] [1]: /lookup/doi/10.1126/science.aac8653

Inhibition of the mammalian target of rapamycin complex 1 signaling pathway reduces itch behaviour in mice.

Activated mammalian target of rapamycin (P-mTOR) has been shown to maintain the sensitivity of subsets of small-diameter primary afferent A-nociceptors. Local or systemic inhibition of the mTOR complex 1 (mTORC1) pathway reduced punctate mechanical and cold sensitivity in neuropathic pain and therefore offered a new approach to chronic pain control. In this study, we have investigated the effects of the rapamycin analog temsirolimus (CCI-779) on itch. Bouts of scratching induced by the histamine-dependent pruritogenic compound 48/80 and histamine-independent pruritogens, chloroquine and SLIGRL-NH2, injected intradermally were significantly reduced by local (intradermal) or systemic (intraperitoneal, i.p.) pretreatment with CCI-779. We also investigated the action of metformin, a drug taken to control type 2 diabetes and recently shown to inhibit mTORC1 in vivo. Although the response to nonhistaminergic stimuli was reduced at all of the time points tested, scratching to compound 48/80 was modified by metformin only when the drug was injected 24 hours before this pruritogen. We also examined the colocalization of P-mTOR with gastrin-releasing peptide, a putative marker for some itch-sensitive primary afferents, and found that P-mTOR was coexpressed in less than 5% of gastrin-releasing peptide-positive fibers in the mouse skin. Taken together, the data highlight the role that P-mTOR-positive A-fibers play in itch signaling and underline the importance of the mTORC1 pathway in the regulation of homeostatic primary afferent functions such as pain and itch. The actions of the antidiabetic drug metformin in ameliorating nonhistamine-mediated itch also suggest a new therapeutic route for the control of this category of pruritus.

Topically Administered Janus-Kinase Inhibitors Tofacitinib and Oclacitinib Display Impressive Antipruritic and Anti-Inflammatory Responses in a Model of Allergic Dermatitis.

The prevalence of allergic skin disorders has increased rapidly, and development of therapeutic agents to alleviate the symptoms are still needed. In this study, we orally or topically administered the Janus kinase (JAK) inhibitors, tofacitinib and oclacitinib, in a mouse model of dermatitis, and compared the efficacy to reduce the itch and inflammatory response. In vitro effects of JAK inhibitors on bone marrow-derived dendritic cells (BMDCs) were analyzed. For the allergic dermatitis model, female BALB/c mice were sensitized and challenged with toluene-2,4-diisocyanate (TDI). Each JAK inhibitor was orally or topically applied 30 minutes before and 4 hours after TDI challenge. After scratching bouts and ear thickness were measured, cytokines were determined in challenged skin and the cells of the draining lymph node were analyzed by means of flow cytometry. In vitro, both JAK inhibitors significantly inhibited cytokine production, migration, and maturation of BMDCs. Mice treated orally with JAK inhibitors showed a significant decrease in scratching behavior; however, ear thickness was not significantly reduced. In contrast, both scratching behavior and ear thickness in the topical treatment group were significantly reduced compared with the vehicle treatment group. However, cytokine production was differentially regulated by the JAK inhibitors, with some cytokines being significantly decreased and some being significantly increased. In conclusion, oral treatment with JAK inhibitors reduced itch behavior dramatically but had only little effect on the inflammatory response, whereas topical treatment improved both itch and inflammatory response. Although the JAK-inhibitory profile differs between both JAK inhibitors in vitro as well as in vivo, the effects have been comparable.

HTR7 Mediates Serotonergic Acute and Chronic Itch

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-regulated 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, including atopic dermatitis. In a mouse model of atopicdermatitis, mice lacking HTR7 or TRPA1 displayed 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.

The construction and implementation of a novel postburn pruritus scale for infants and children aged five years or less: introducing the Toronto Pediatric Itch Scale.

The authors' objectives were to design, refine, validate and implement a behavior-anchored postburn pruritus scale for children aged 5 years or less. We engaged a range of professionals involved in the care of children with burns. We used Q-methodology in interprofessional team exercises to identify and stratify itch behaviors into categories of increasing severity, and then iteratively refined these into a draft scale. We used a range of quantitative and qualitative techniques to assess the utility, feasibility, and validity of the scale and refined it accordingly. During the implementation phase we collected some preliminary reliability data. We generated a 4-point scale of itch severity with simple descriptors of each score. We also designed a separate guidance note and example behaviors that could be used to orientate new users without the need for rater training. End-user interviews revealed high levels of feasibility and content validity. The reliability data showed moderate inter-observer agreement, with a Cohen's kappa of 0.52 (P < .001). We have developed and implemented a behavioral post-burn pruritus scale for use in children aged less than 5 years and have demonstrated its utility, feasibility, validity, and reliability. The development of a validated symptom scoring scales will allow for the conduct of high-quality quantitative clinical trials and the subsequent implementation of evidence-based management protocols.

The Epithelial Cell-Derived Atopic Dermatitis Cytokine TSLP Activates Neurons to Induce Itch

Atopic dermatitis (AD) is a chronic itch and inflammatory disorder of the skin that affects one in ten people. Patients suffering from severe AD eventually progress to develop asthma and allergic rhinitis, in a process known as the "atopic march." Signaling between epithelial cells and innate immune cells via the cytokine thymic stromal lymphopoietin (TSLP) is thought to drive AD and the atopic march. Here, we report that epithelial cells directly communicate to cutaneous sensory neurons via TSLP to promote itch. We identify the ORAI1/NFAT calcium signaling pathway as an essential regulator of TSLP release from keratinocytes, the primary epithelial cells of the skin. TSLP then acts directly on a subset of TRPA1-positive sensory neurons to trigger robust itch behaviors. Our results support a model whereby calcium-dependent TSLP release by keratinocytes activates both primary afferent neurons and immune cells to promote inflammatory responses in the skin and airways.

P44 Does the slow-releasing hydrogen sulfide donor GYY4137 controls inflammation and pruritogen responses in mouse skin?

A growing body of evidence suggests that hydrogen sulphide (H2S) plays a crucial role in both physiological and pathophysiological process, including heart failure, inflammatory lung and articular diseases, erectile dysfunction, diabetes, among others. Poor diabetes control in women (mean glycosylated hemoglobin level The aim of this study is to determine the effectiveness of the slow H2S-releasing donor GYY4137 in reducing itch behaviour and cutaneous inflammation in mouse dorsal skin. The animal protocols were approved by the local ICB-USP Ethics Committee. In order to produce itching behaviour, male BALB/c mice (25–30 g) were anaesthetised with isoflurano and i.d. injected with histamine (1 μmol site−1) alone or co-injected with GYY4137 (0.3–30 nmol site−1; 0.05 ml). Itching behaviour was estimated by measuring the number of scratching bouts over a 40 min period. In a separate set of experiments, animals were pre-treated (−40 min) with the ATP-sensitive K+ (KATP) channel blocker glibenclamide, and then i.d. injected with histamine and Na2S (3 nmol site−1). In addition, we investigated the presence of mRNA for the enzymes cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfur transferase (3-MST) in the mouse naive skin via qualitative RT-PCR. Data are presented as mean ± SEM and were analyzed by ANOVA followed by the Dunnett test. Values of P The i.d. injection of histamine significantly increased the itching frequency as compared to its vehicle Tyrode. Increasing doses of GYY4137 (1–10 nmol site−1) dose-dependently reduced histamine-induced pruritus but failed to inhibit plasma extravasation evoked by histamine. Suppression of histamine-induced pruritus by the H2S donor was not reversed by glibenclamide. A marked mRNA expression for CBS and 3-MST, but not for CSE, was seen in the mouse naive skin. These findings indicate that low doses of GYY4137 in the skin decrease histamine-induced pruritus without affecting the associated local inflammation. This protective effect does not appear to be mediated by KATP channels activation. Financial support: FAPESP, CAPES and CNPq.

Assessment of Pain and Itch Behavior in a Mouse Model of Neurofibromatosis Type 1

Neurofibromatosis type 1 (NF1) is characterized primarily by tumor formation in the nervous system, but patients report other neurological complications including pain and itch. Individuals with NF1 harbor 1 mutated NF1 allele causing heterozygous expression in all of their cells. In mice, Nf1 heterozygosity leads to hyperexcitability of sensory neurons and hyperproliferation of mast cells, both of which could lead to increased hypersensitivity and scratching in response to noxious and pruritic stimuli. To determine whether Nf1 heterozygosity may increase pain and itch behaviors independent of secondary effects of tumor formation, we used mice with a targeted, heterozygous Nf1 gene deletion (Nf1±) that lack tumors. Nf1± mice exhibited normal baseline responses to thermal and mechanical stimuli. Moreover, similar to wild-type littermates, Nf1± mice developed inflammation-induced heat and mechanical hypersensitivity, capsaicin-induced nocifensive behavior, histamine-dependent or -independent scratching, and chronic constriction injury-induced cold allodynia. However, Nf1± mice exhibited an attenuated first phase of formalin-induced spontaneous behavior and expedited resolution of formalin-induced heat hypersensitivity. These results are not consistent with the hypothesis that Nf1 heterozygosity alone is sufficient to increase pain and itch sensation in mice, and they suggest that additional mechanisms may underlie reports of increased pain and itch in NF1 patients. PerspectiveThis study assessed whether Nf1 heterozygosity in mice increased hypersensitivity and scratching following noxious and pruritic stimuli. Using Nf1± mice lacking tumors, this study finds no increases in pain or itch behavior, suggesting that there is no predisposition for either clinical symptom solely due to Nf1 heterozygosity.

Excitatory Superficial Dorsal Horn Interneurons Are Functionally Heterogeneous and Required for the Full Behavioral Expression of Pain and Itch

To what extent dorsal horn interneurons contribute to the modality specific processing of pain and itch messages is not known. Here, we report that loxp/cre-mediated CNS deletion of TR4, a testicular orphan nuclear receptor, results in loss of many excitatory interneurons in the superficial dorsal horn but preservation of primary afferents and spinal projection neurons. The interneuron loss is associated with a near complete absence of supraspinally integrated pain and itch behaviors, elevated mechanical withdrawal thresholds and loss of nerve injury-induced mechanical hypersensitivity, but reflex responsiveness to noxious heat, nerve injury-induced heat hypersensitivity, and tissue injury-induced heat and mechanical hypersensitivity are intact. We conclude that different subsets of dorsal horn excitatory interneurons contribute to tissue and nerve injury-induced heat and mechanical pain and that the full expression of supraspinally mediated pain and itch behaviors cannot be generated solely by nociceptor and pruritoceptor activation of projection neurons; concurrent activation of excitatory interneurons is essential.

Itch-specific neurons

Dorsal root ganglia (DRG) neurons that respond to itchy stimuli also respond to painful stimuli, raising the question of whether any DRG neurons specifically mediate itch. The authors showed that MAS-related G protein-coupled receptor A3 (MRGPRA3)-expressing (MRGPRA3+) DRG neurons selectively innervate the epidermis and respond to both itchy and noxious stimuli. Mrgpra3 ablation in mice reduced itch behaviour but not pain behaviour, and selective activation of MRGPRA3+ neurons with a noxious stimulus induced itch behaviour but not pain behaviour, suggesting that MRGPRA3+ DRG neurons specifically mediate itch.

Psychophysiological Effects of Stress Management in Patients with Atopic Dermatitis: A Randomized Controlled Trial

Atopic dermatitis leads to, and can be triggered by, stress. Psychological interventions have been shown to have positive effects on skin status, itch and scratching behaviour. However, it has not been analysed whether stress management leads to a change in physiological stress level and psychophysiological stress reaction under acute stress in this patient group. In this study 28 patients with atopic dermatitis were randomized to an experimental group (cognitive behavioural stress management) or a control group. The endocrine stress level and skin status were measured before and after the stress management programme. A public-speaking paradigm was used to induce acute stress. The study revealed that the experimental group had a tentatively reduced cortisol awakening response after the stress management programme. In addition, the experimental group remained calmer and showed lower salivary cortisol levels under acute stress. Thus, stress management might be a useful addition to standard treatment in patients with atopic dermatitis.

Role of nociceptive arcuate nucleus neurons in chloroquine-induced pruritic behaviors in mice

Despite its clinical importance, the underlying central mechanisms of pruritic behaviors are poorly understood. To investigate the role of nociceptive arcuate nucleus neurons in chloroquine-induced pruritic behaviors in mice, we tested the effect of arcuate nucleus neurons and interscapular brown adipose tissue (IBAT) on itch produced by intradermal injection of chloroquine in the nape of the neck. Our results provide several lines of evidence for an important role of nociceptive arcuate nucleus neurons in chloroquine-induced pruritic behavior: (1) Intradermal microinjection of chloroquine resulted in a dramatic increase in itch behaviors accompanied by the activation of c-Fos positive neurons in arcuate nucleus; (2) Microinjection of chloroquine significantly increased IBAT temperature in the mice. These findings suggested that chloroquine-induced pruritic behaviors were associated with the activity of nociceptive arcuate nucleus neurons.

Supplementation of the fermented soy product ImmuBalance™ effectively reduces itching behavior of atopic NC/Tnd mice

Effects of probiotics on the prevention of atopic diseases have been proposed recently. Although we have already reported the suppressive effects of the probiotic, ImmuBalance™, on a mouse model for peanuts allergy, its influence on atopic diseases remains unclear. Potential efficacy of ImmuBalance™, which is the fermented soy product, on treatment of atopic dermatitis (AD) was investigated using a mouse model for human AD, NC/Tnd mice. For in vivo study, ImmuBalance containing chow or a control diet were fed to NC/Tnd mice with moderate dermatitis for 2 weeks. Topical application of FK506 ointment was used as a positive control. Clinical skin severity scores, scratching behaviors, trans-epidermal water loss (TEWL), and histological features were analyzed. For in vitro study, suppressive effect of ImmuBalance™ on nerve growth factor (NGF)-activated neurite outgrowth of PC12 cells was examined. Clinical skin severity scores of the mice fed with ImmuBalance containing chow were gradually reduced as well as the mice treated with FK506. Feeding with ImmuBalance completely inhibited the increase in scratching behavior of NC/Tnd mice. The value of TEWL of NC/Tnd mice fed with ImmuBalance was significantly decreased. In addition, histological examination revealed that application of ImmuBalance decreased the number of PGP9.5-positive neuronal fibers in the lesional skin. When ImmuBalance extract was added to the culture, NGF-activated neurite outgrowth of PC12 cells was diminished through the inhibition of the phosphatidylinositol 3-kinase phosphorylation. ImmuBalance could exhibit favorable alterations on AD symptoms, particularly through down regulation of the itch sensation.

VGLUT2-Dependent Sensory Neurons in the TRPV1 Population Regulate Pain and Itch

The natural response to itch sensation is to scratch, which relieves the itch through an unknown mechanism. Interaction between pain and itch has been frequently demonstrated, and the selectivity hypothesis of itch, based on data from electrophysiological and behavioral experiments, postulates the existence of primary pain afferents capable of repressing itch. Here, we demonstrate that deletion of vesicular glutamate transporter (VGLUT) 2 in a subpopulation of neurons partly overlapping with the vanilloid receptor (TRPV1) primary afferents resulted in a dramatic increase in itch behavior accompanied by a reduced responsiveness to thermal pain. The increased itch behavior was reduced by administration of antihistaminergic drugs and by genetic deletion of the gastrin-releasing peptide receptor, demonstrating a dependence on VGLUT2 to maintain normal levels of both histaminergic and nonhistaminergic itch. This study establishes that VGLUT2 is a major player in TRPV1 thermal nociception and also serves to regulate a normal itch response.

An Itch To Be Scratched

The description of itch (formally known as pruritus) as an "unpleasant sensation that elicits the desire or reflex to scratch" (Ikoma et al., 2006) is immediately familiar. Research in the field of pruritoception has added to our understanding of this area of sensory neurobiology as it pertains to both normal and pathological conditions. In particular, much progress has been made on the mechanisms and circuits of itch, which we review here.