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Su2037

Role of Neuronal and Non-Neuronal Transient Receptor Potential VanilloidType 1 and Sensory Neuropeptides in DSS-Induced Peripheral InflammatoryChangesKenjiro Matsumoto, Takuji Hosoya, Kenta Kijiyama, Rikako Suzuki, Kimihito Tashima,Toshihiko Murayama, Syunji Horie

BACKGROUND & AIMS: The activation of transient receptor potential vanilloid type 1(TRPV1) leads to release of sensory neuropeptides such as substance P (SP), neurokinin A(NKA) and calcitonin gene-related peptide (CGRP) from the nerve endings, and theseneuropeptides in turn initiate the biochemical cascade known as neurogenic inflammation.TRPV1 channels and the sensory neuropeptides are expressed not only in sensory nervefibers but also in non-neuronal cells in the mucosa of large intestine. The aim of this studywas to investigate alteration of the sensory neuropeptides and TRPV1 channels in mucosaof DSS-induced colitis mice and to characterize their non-neuronal cells. METHODS: Colitiswas induced by 3% dextran sulfate sodium (DSS) solution given as drinking water for 7days in C57BL/6 mice. To evaluate visceral nociception, colorectal distension was performedin mice treated with vehicle or BCTC on day 7 of DSS treatment. Immunohistochemicalanalysis was performed using rectal tissues of mice. SP, NKA, CGRP, 5-HT, F4/80, keratin,TNF-α, CD11c, and CD4 were detected by indirect staining with their specific antibodies.TRPV1-immunoreactivity was detected by using immunohistochemical staining with fluor-escein-conjugated tyramide amplification. RESULTS: TRPV1 antagonist BCTC significantlyattenuated the visceral hyperalgesia to control level in DSS-induced colitis model. Wecompared the number of the nerve fibers and non-neuronal cells expressing TRPV1 channelsand sensory neuropeptides in normal and DSS-induced colitis model mice. The number ofTRPV1- and CGRP-expressing nerve fibers is significantly increased in colitis model. Thenon-neuronal TRPV1 cells were markedly increased but non-neuronal CGRP-expressingcells were not observed in colitis model. No significant alteration in the number of SP andNKA positive nerve fibers was detected. On the other hand, SP and NKA positive cells weredrastically increased in colitis model. Next, double labeling studies were carried out tocharacterize TRPV1-, SP-, and NKA-expressing cells under inflammatory state. We foundtwo types of non-neuronal TRPV1-expressing cells in mucosa of colitis model. One is keratinpositive cells located upper part of mucosa. Another is TNF-α and/or F4/80 positive cellslocated middle part of the mucosa. Both types of TRPV1-expressing cells did not colocalizewith CD4 and CD11c. SP and NKA positive cells almost completely colocalized with 5-HTin experimental colitis model mice. CONCLUSION: These results suggest that TRPV1-immunopositive epithelial cells and macrophage are associated with visceral hyperalgesiain colitis.

S-540AGA Abstracts

Su2038

Mechano- and Chemo-Sensitive Spinal Dorsal Root Ganglia (DRG) AfferentNerve Fibers in the Esophageal MuscleFei Ru, Peter Banovcin, Marian Kollarik

It is often assumed that the sensory transduction sites of the esophageal afferent nerve fibersresponsive tomucosal touch but unresponsive to esophageal distention are located exclusivelyin the mucosa (mucosal afferent nerve fibers). We addressed the hypothesis that the eso-phageal spinal DRG nociceptors have mechano- and chemo- transduction sites in the eso-phageal muscle. Ex vivo extracellular single unit recordings were made from the T2-T3 DRGneurons projecting afferent nerve fibers into the esophagus in the isolated innervated guineapig esophagus preparation. Virtually all DRG fibers found by using an electrical stimulusresponded to focal mechanical stimulation (focal compression 91/94; focal touch with vonFrey filaments, 20/22) and capsaicin (1microM, 27/29). We focused on the afferent fibersunresponsive to esophageal distention that comprised the majority of the DRG fibers (approx-imately 80% , 42/54 DRG fibers failed to respond to esophageal distention to 100mmHg).Removal of the esophageal mucosa did not abolish the response to von Frey filaments (n=15) and did not appreciably change the mechanical threshold (1.5±0.4 -fold increase, p=0.3, n=5). In 6/6 experiments further removal of the serosal layer had no effect on themechanical response. The response of DRG fibers to capsaicin (1microM) was not appreciablydifferent in the preparations with the mucosa intact and removed (peak discharge 8.6±1.4Hz,n=14 vs. 10.9±1.4Hz, n=10, respectively, p>0.3). In the preparations with the mucosaremoved acid (pH=6.5-5.5) was also effective to evoke action potential discharge in 5/6DRG fibers. In the preparations with the mucosa removed detailed analysis by using vonFrey filaments revealed relatively large (axial length 25±2 mm, n=7) mechanosensitivereceptive fields sometimes consisting of multiple separated mechanosensitive spots. Consist-ent with this observation, branching eGFP-expressing fibers located in the esophageal musclewere visualized following the transfection of thoracic DRG neurons with the adeno-associatedvirus vector encoding enhanced green fluorescent protein (eGFP). We conclude that thedistention-insensitive esophageal DRG nociceptive fibers have extensive sensory receptivefields in the esophageal muscle. While published anatomical studies show that the spinalDRG fibers project also into esophageal mucosa the transduction properties of these projec-tions remain to be functionally determined. Supported by NIH DK074480 (M.K.), AstraZ-eneca IRUS33550004 (M.K.) and VEGA 1/0276/10 (P.B.)

Su2039

Dysfunctional Endogenous Pain Modulation (EPM): A Surrogate Biomarker inIrritable Bowel Syndromes (IBS)?Yang Cao, Xinhua Li, Reuben K. Wong, Khek Yu Ho, Clive H. Wilder-Smith

INTRODUCTION: endogenous pain modulation is a central homeostatic control mechanismclosely linked to other non-sensory homeostatic centres. Measures of EPM have in a fewrecent studies been shown to correlate with clinical measures of somatic and neuropathicpain as well as pain relief, suggesting their potential usefulness as surrogate markers forclinical pain outcomes. Sensory hypersensitivity and abnormal EPM have been demonstratedin IBS by sensory testing with heterotopic stimulation and functional brain imaging studies,but correlations between EPM and clinical pain and other symptoms in IBS have not beenreported. AIMS &METHODS: We studied correlations between clinical IBS symptom ratingsand EPM induced by standard heterotopic stimulation in 12 IBS patients (Rome III, 7females). Foot heat and rectal distension sensation and pain thresholds were determinedusing an ascending method of limits. Subsequently, moderate foot and rectal pain individuallytitrated to an intensity of 30-60 on a VAS of 0-100 were applied separately and together(heterotopic stimulation) six times for 30 seconds in randomised sequence during brainfunctional Magnetic Resonance Imaging (fMRI). The following were recorded during theweek before sensory testing: IBS symptom severity (IBS-SSS questionnaire) and averageabdominal pain and discomfort by VAS. Correlations were analyzed by linear regression.RESULTS: The magnitude of EPM correlated with IBS-related abdominal pain intensity (r=0.64, p<0.02), abdominal discomfort intensity (r=0.69, p<0.01), quality of life scores (r=0.71, p<0.01), as well as IBS-SSS scores (r=0.84, p<0.001). The duration of IBS pain showedno significant association with EPM. CONCLUSIONS: This data suggests there is a relevantcorrelation between the magnitude of EPM and clinical pain and symptoms in IBS patients.If confirmed in an ongoing study with a larger group of patients with treatment, EPMmeasurementmay become a useful surrogate biomarker in pathophysiological and therapeuticresearch of IBS. The current study was support by Singapore National Medical ResearchCouncil, Individual Research Grant.

Su2040

Repeated Psychological Stress Induces an Immediate Naloxone-IndependentVisceral Analgesia to Colorectal Distension in RatsMuriel H. Larauche, Agata Mulak, Yong Sung Kim, Mulugeta Million, Yvette Tache

Background: A variety of stressful stimuli have been shown to elicit analgesia - a phenomenonknown as stress-induced analgesia (SIA). Both opioid and non-opioid forms of SIA havebeen identified and found to depend on the type of stressor and its duration. While in naïveanimals exposed to psychological stress opioids have been shown to participate in somaticSIA, in rats previously exposed to pain endogenous opioids can paradoxically be associatedwith the development of stress-induced hyperalgesia via the recruitment of NMDA-dependentpronociceptive systems (Rivat et al., Neuropsychopharm 2007). Aims: To characterizewhether the commonly used water avoidance stress (WAS) induces a naloxone sensitive orindependent visceral analgesia as monitored by the visceromotor response (VMR) to colorectaldistension (CRD) in female and male rats. Methods: Male and female Wistar rats (7-8 weeks,2-4/cage) were monitored for VMR to graded phasic CRD (10, 20, 40, 60 mmHg, 20 secduration, 4 min interstimulus intervals) using a newly developed manometric technique(Larauche et al., AJP 2009). The 1st CRD (day 0) served as baseline VMR, then rats wereexposed to 4 days of WAS (1h/day). Two groups were injected subcutaneously (sc) withnaloxone (1 mg/kg) (n=9) or saline (0.3 ml) (n=9) 10 min before each WAS, another control