Yasushi Kuraishi

Gastrin-releasing peptide induces itch-related responses through mast cell degranulation in mice.

Gastrin-releasing peptide (GRP), secreted from the central terminals of primary afferents, is involved in the transmission of itch signals in the spinal dorsal horn. Although primary afferents containing GRP are distributed throughout the skin, the role of peripherally released GRP in the itch response is unknown. We investigated whether GRP acts on the skin to induce an itch response in mice. Intradermal injections of GRP(18-27) (1-300 nmol/site) elicited scratching. GRP(18-27)-induced scratching was inhibited by the μ-opioid receptor antagonist naltrexone hydrochloride, the BB(2) bombesin receptor antagonist RC-3095, the H(1) histamine receptor antagonists fexofenadine hydrochloride and chlorpheniramine maleate, and the PAR(2) proteinase-activated receptor antagonist FSLLRY-NH(2). Mast cell deficiency significantly, but not completely, reduced the GRP(18-27)-induced scratching. BB(2) bombesin receptors are present in mast cells in the skin, and intradermal injection of GRP(18-27), not only induced scratching, but also led to mast cell degranulation. GRP(18-27)-induced mast cell degranulation was inhibited by the BB(2) bombesin receptor antagonist RC-3095. These results suggest that peripherally released GRP can induce an itch response, at least partly, through activation of BB(2) receptors present in the mast cells, triggering their degradation and the release of histamine and the serine proteinase, tryptase.

Noradrenergic regulation of itch transmission in the spinal cord mediated by α-adrenoceptors.

It has recently been shown that clonidine suppresses itch-related responses via its action on α(2)-adrenoceptors in the spinal cord, raising the possibility that the descending noradrenergic system regulates itch signaling in the spinal cord. In this study, we investigated whether the transmission of itch signals in the spinal cord is under tonic inhibition by the descending noradrenergic system. An intraplantar injection of serotonin in mice induced biting of the treated paw (an itch-related response). An intrathecal injection of 6-hydroxydopamine (catecholaminergic neurotoxin) enhanced the itch-related response. There was a significant inverse correlation between the response and noradrenaline content. An intrathecal injection of phentolamine (α-adrenoceptor antagonist) enhanced serotonin-induced biting, although prazosin (α(1)-, α(2B)-, and α(2C)-adrenoceptor antagonist) and yohimbine (α(2)-adrenoceptor antagonist) had no effects. Intrathecal injections of phenylephrine (α(1)-adrenoceptor agonist) and clonidine (α(2)-adrenoceptor agonist) inhibited serotonin-induced biting. The action of phenylephrine was antagonized by intrathecal prazosin but not 5-methylurapidil (α(1A)-adrenoceptor antagonist), cyclazosin (α(1B)-adrenoceptor antagonist), and BMY 7378 (α(1D)-adrenoceptor antagonist). mRNAs encoding α(1A)-, α(1B)-, α(2A)-, α(2B)-, and α(2C)-adrenoceptor subtypes were expressed in the dorsal root ganglion and spinal dorsal horn. These results suggest that the descending noradrenergic system exerts tonic inhibition on itch signaling in the spinal cord. Both α(1)- and α(2)-adrenoceptors may be involved in the tonic inhibition of itch signaling and the stimulation of either α-adrenoceptor subtype may result in the inhibition of itch.

Cathepsin E induces itch-related response through the production of endothelin-1 in mice.

This study investigated the pruritogenic potency of cathepsin E, an aspartic protease, and its mechanisms in mice. An intradermal injection of cathepsin E to the rostral back elicited scratching, an itch-associated response, of the injection site. This action was inhibited by the aspartic protease inhibitor pepstatin A, the endothelin ET(A) receptor antagonist BQ-123, and the opioid receptor antagonists naltrexone and naloxone, but not by the H(1) histamine receptor antagonist terfenadine, the proteinase-activated receptor-2 antagonist FSLLRY-NH(2), or mast cell deficiency. Pepstatin A inhibited scratching induced by intradermal injection of the mast-cell degranulator compound 48/80, but not by tryptase, a mast-cell mediator. An intradermal injection of cathepsin E increased endothelin-1 levels in the skin at the injection site. Preproendothelin-1 mRNA was present in primary cultures of keratinocytes, and immunohistochemistry using an antibody recognizing endothelin-1 and big-endothelin-1 revealed immunoreactivity in the epidermis, especially in the prickle and granular cell layers, but not in the basal cell layer. These results suggest that cathepsin E is an endogenous itch inducer, and that its action is mediated at least in part by the production of endothelin-1 in the epidermis.

Involvement of serine protease and proteinase-activated receptor 2 in dermatophyte-associated itch in mice

We investigated the involvement of serine protease and proteinase-activated receptor 2 (PAR2) in dermatophyte-induced itch in mice. An intradermal injection of an extract of the dermatophyte Arthroderma vanbreuseghemii (ADV) induced hind-paw scratching, an itch-related behavior. ADV extract-induced scratching was inhibited by the opioid receptor antagonists naloxone and naltrexone, the serine protease inhibitor nafamostat mesylate, and the PAR2 receptor antagonist FSLLRY-NH2. ADV extract-induced scratching was not inhibited by the H1 histamine receptor antagonist terfenadine or by mast cell deficiency. Heat pretreatment of the ADV extract markedly reduced the scratch-inducing and serine protease activities. Proteolytic cleavage within the extracellular N terminus of the PAR2 receptor exposes a sequence that serves as a tethered ligand for the receptor. The ADV extract as well as tryptase and trypsin cleaved a synthetic N-terminal peptide of the PAR2 receptor. The present results suggest that serine protease secreted by dermatophytes causes itching through activation of the PAR2 receptors, which may be a causal mechanism of dernatophytosis itch.

Clonidine inhibits itch-related response through stimulation of α2-adrenoceptors in the spinal cord in mice

The present study investigated whether clonidine - an α(2)-adrenoceptor agonist known to relieve pain - is able to suppress itch-related behavior in mice. An intraplantar injection of serotonin induced biting (an itch-related response), which was inhibited by intraperitoneal and intrathecal, but not intraplantar or intracisternal, clonidine injections. The effect of intrathecal clonidine was inhibited by intrathecal injections of phentolamine (a non-selective α-adrenoceptor antagonist) and yohimbine (a selective α(2)-adrenoceptor antagonist), but not by prazosin (a selective α(1)-adrenoceptor antagonist). The effect of intraperitoneal clonidine was also inhibited by intrathecal yohimbine. These results suggest that clonidine is an effective antipruritic agent and that the effect is mainly mediated by the stimulation of α(2)-adrenoceptors in the dorsal horn.

Involvement of leukotriene B4 in dermatophyte-related itch in mice

BACKGROUNDnProteinase-activated receptor-2 (PAR2) is involved in dermatophyte-induced scratching and leukotriene B4 (LTB4) release from keratinocytes. We investigated whether PAR2-mediated LTB4 production is involved in dermatophyte-induced scratching.nnnMETHODSnDermatophyte extract was injected intradermally and scratching was observed in mice. LTB4 was determined by enzyme immunoassay.nnnRESULTSnDermatophyte extract-induced scratching was inhibited by zileuton (5-lipoxygenase inhibitor), ONO-4057 (LTB4 antagonist), FSLLRY-NH2 (PAR2 antagonist), and anti-PAR2 antibody. Dermatophyte extract injection increased the cutaneous content of LTB4, which was inhibited by zileuton and FSLLRY-NH2.nnnCONCLUSIONnThese results suggest the involvement of LTB4 in dermatophyte-associated itch. LTB4 production might be due to PAR2 stimulation in the skin.

BB2 bombesin receptor-expressing spinal neurons transmit herpes-associated itch by BB2 receptor-independent signaling.

Although spinal neurons expressing BB2 bombesin receptors are suggested to be involved in itch transmission, their role in pathological itch remains unknown. Because itch is often observed in patients with herpes zoster, we examined the role of BB2 receptor-expressing spinal neurons in herpes-associated itch in mice. Transdermal inoculation of human herpes virus 1 on the midflank produced herpes zoster-like skin lesions and caused the mice to scratch (itch-related behavior) and lick (pain-related behavior) the affected skin. Ablation of BB2 receptor-expressing spinal neurons by intrathecal treatment with a bombesin–saporin conjugate decreased the scratching but not the licking. Intrathecal administration of the BB2 receptor antagonist Leu13-&psgr;-(CH2NH)Leu14-bombesin decreased BB2 receptor agonist GRP18–27-induced scratching in naive mice but not herpes-associated scratching. The present results suggest that BB2 receptor-expressing spinal neurons transmit herpes-associated itch by BB2 receptor-independent signaling.

Peripheral mechanisms of the induction of itch

s / Neuroscience Research 68S (2010) e4–e52 e49 S3-9-1-3 How is glycine cleavage system involved in cognitive dysfunction of schizophrenia? Chihiro Kakiuchi Department of Neuropsychiatry, University of Tokyo Glutamatergic dysfunction, especially NMDA receptor hypofunction, has been shown to be involved in the pathophysiology of schizophrenia. Glycine or d-serine enhances NMDA receptor activation through glycine binding site, and both molecules are suggested to be effective for the symptoms of the illness. Glycine cleavage system(GCS), known as a causative pathway of glycine encephalopathy(GE)/hyperglycinemia, is an enzyme complex composed of GCSP, GCST, GCSL and GCSH, which breaks down glycine to carbon dioxide and ammonia. Interestingly, d-serine is decreased in the brain of GE, and recently, co-localization of this system with NMDA receptor has been reported. Our preliminary genetic study suggested the association of this system with schizophrenia. Possible role of GCS in the pathophysiology of schizophrenia including cognitive dysfunction will be discussed. doi:10.1016/j.neures.2010.07.462 S3-9-1-4 Use of neurophysiological biomarkers for understanding the pathways from genes to real-world daily functioning in schizophrenia Gregory A. Light , Tiffany Greenwood, Joyce Sprock, Neal Swerdlow, David Braff Department of Psychiatry, 0804, University of California, San Diego Schizophrenia patients have widespread deficits ranging from abnormalities in sensory processing to impairments in cognition and daily living. Mismatch negativity (MMN) is a neurophysiological biomarker observed in the EEG that probes the earliest stages of sensory information processing and can be elicited in the absence of directed attention. To assess the genetic architecture of MMN, a large-scale candidate gene analysis was performed on 203 schizophrenia patients and 119 controls from the UCSD Schizophrenia Program using the Consortium on the Genetics of Schizophrenia (COGS) custom SNP chip. The single marker analyses in the Caucasian subset revealed significant associations with 25 genes (all empirical p<0.01). These genes included BDNF, GRIN3A, DISC1, CTNNA2, NRG1, and ERBB4. Several of these genes also interact molecularly (e.g., NRG1 and ERBB4). Schizophrenia patients also had significantly reduced MMN (p<0.01). MMN deficits were associated with impaired performance on tests of working memory (p<0.01), verbal recall (p<0.01), and more severe levels of negative symptoms (p<0.01). MMN deficits were also associated with reduced performance (p<0.05) on a comprehensive functional skills assessment battery (e.g., ability to perform basic financial tasks), and significantly (p<0.01) lower ratings on several measures of functional status (e.g., independence in living situation, managing finances, Scale of Functioning, Global Assessment of Functioning Scale). In contrast, MMN was not associated with performance on other cognitive measures or positive symptoms. Results from structural equation modeling will also be presented to demonstrate the pathways from MMN and related neurophysiological biomarkers (e.g., N100, P3a) to measures of real-world daily functioning. doi:10.1016/j.neures.2010.07.463 S3-9-1-5 How can schizophrenic cognitive dysfunction be integrated with experimental data? Ichiro Sora Department of Biological Psychiatry, Tohoku University Graduate School of Medicine The hyperdopaminergia of dopamine transporter knockout (DAT KO) mice, judged in terms of extracellular dopamine levels in the striatum or dopamine-associated behaviors such as hyperactivity, have led to the suggestion that DAT KO mice can be used as animal models of schizophrenia. Pre-pulse inhibition (PPI) deficits are observed in several psychiatric disorders, especially schizophrenia. This impairment of sensorimotor gating has been postulated to reflect at least some portion of the cognitive dysfunction observed in schizophrenia. DAT KO mice have deficits in PPI of the acoustic startle reflex, a model of sensorimotor gating which are also reversed by treatment with a number of psychostimulants. The underlying deficit in DAT KO mice is likely to involve alterations in the balance between ventral striatal and prefrontocortical activity. In part, these results from an oddity of dopamine function in the prefrontal cortex whereby uptake is normally mediated by norepinephrine transporter (NET) rather than DAT. One consequence of this situation is that in the absence of DAT in DAT KO mice there are profound alterations in extracellular dopamine concentrations in the ventral striatum, while the prefrontal cortex remains substantially unaffected, thereby potentially altering the balance of activity between the prefrontal cortex and the ventral striatum. This would appear to have dramatic effects on responses to psychostimulants, which impairs PPI in wild-type mice, but normalizes PPI in DAT KO mice. This study went on to show that, prefrontal cortical NET blockade and consequent enhancement of prefrontal cortical extracellular dopamine mediates the reversal of PPI deficits in DAT KO mice. doi:10.1016/j.neures.2010.07.464 S3-9-2-1 Peripheral mechanisms of the induction of itch Tsugunobu Andoh , Yasushi Kuraishi University of Toyama Itch is a skin sensation that provokes a desire to scratch. Although histamine, a mast cell mediator, has been thought to play an essential role in itch, many pruritic diseases, such as atopic dermatitis, except acute urticaria respond poorly to H1 histamine receptor antagonists. However, recent development of the evaluation methods of itch in animal and animal models of itch has been shedding light on the mechanisms and new peripheral mediators of itch. Many endogenous itch mediators have been revealed, but this talk deals with protease and lipid itch mediators. Proteases have long been known to elicit itch in humans and recent works have shown that serine proteinase, such as tryptase, another mast cell mediator, causes itch mainly through the action on PAR2 proteinase-activated receptor and is involved in pruritus of humans with atopic dermatitis and mice with chronic allergic dermatitis. Regarding lipid itch mediators,arachidonic acid metabolites, such as prostaglandin E2, thromboxane A2, and leukotriene B4, mainly produced by keratinocytes induce and/or enhance itch. In the healthy skin, sphingomyelin is hydrolyzed to ceramide, which is important to keep skin hydration. In atopic dermatitis, sphingomyelin is highly catalyzed to sphingosylphosphorylcholine and ceramide is decreased. Sphingosylphosphorylcholine acts on primary afferent to produce itch and it also acts on keratinocytes to produce leukotriene B4. Thus, abnormal metabolism of sphingomyelin may be a cause of pruritus and skin dryness of atopic dermatitis. Peripheral mechanisms of pruritus are not simple but rather many endogenous mediators may play roles in pruritus in consort. doi:10.1016/j.neures.2010.07.465 S3-9-2-2 Intractable itch and nerve fibers-penetration mechanisms of nerve fibers into epidermis Mitsutoshi Tominaga 1 , Kenji Takamori 1,2 1 Institute for Environ and Gender Specific Med, Juntendo Univ Graduate School of Med, Urayasu, Japan 2 Dept Dermatol, Juntendo Univ Urayasu Hospital, Urayasu, Japan Itch (or pruritus) is an unpleasant sensation inducing a desire to scratch. As part of the body’s defense mechanism, itch guards the skin against potentially dangerous organisms or stimuli. Moreover, itch is a major and distressing symptom of many skin and systemic diseases. Histamine is the best-known pruritogen in humans and is also regarded as an experimental itch-causing substance. Clinically, antihistamines, i.e., H1-receptor blockers, are commonly used to treat all types of itch resulting from renal and liver diseases, as well as from serious skin diseases, such as atopic dermatitis. However, antihistamines often lack efficacy in patients with chronic itch, as there are many other itch-causing substances, such as proteases, neuropeptides, cytokines, and opioids, and their cognate receptors, such as thermoreceptors, PAR-2, and opioid receptors. Itch is transmitted to the central nervous system by specialized nerve fibers and sensory receptors. Recent studies regarding gastrin-releasing peptide (GRP) receptors indicated that itch and pain have their own neuronal pathways. As a cutaneous sensory perception, itch may require excitation of neuropeptide-containing free nerve endings of unmyelinated C-fibers, such as GRP+ fibers. In addition, neuronal sensitization caused by activation of itch-related receptors on sensory nerve fibers and increases in the number of these fibers, as well as neurogenic inflammation are partly involved in chronic itch. In this symposium, we focus on epidermal nerve fibers that are partly involved in the regulation of itch, and discuss the penetration mechanisms of nerve fibers into the epidermis of pruitic skin such as atopic dermatitis. doi:10.1016/j.neures.2010.07.466