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Tu2124 P2X7 Receptor Modulate Primary Afferent Responses to Colorectal Distension in Rat Models With Visceral Pain
Tu2122 Healthy Volunteers With the Pain Protective GTP Cyclohydrolase-1 Haplotype Who Sensitise to Oesophageal Acidification Possess Significantly Higher Pain Thresholds David C. Bulmer, Claude A. Botha, Wendy Winchester, L. Ashley Blackshaw, Charles H. Knowles, Qasim Aziz
Tu2120 Quality, Intensity and Location of the Urge to Defaecate Differs Between Patients With Constipation and Healthy Subjects Rebecca Burgell, Michal M. Szczesniak, Priyanka Yerragorla, Emma V. Carrington, Sahar Mohammed, Peter J. Lunniss, Philip G. Dinning, S. Mark Scott
Introduction: GTP Cyclohydrolase-1 (GCH-1) is an enzyme responsible for the de novo synthesis of tetra borohydrate (BH4), a cofactor in monoamine and nitric oxide synthesis. Genetic studies have identified the presence of a pain protective GCH-1 haplotype which is associated with reduced BH4 production and lower pain scores in chronic pain patients or healthy volunteers following algogenic stimuli. The aim of this study was to examine influence of GCH-1 haplotype in healthy volunteers characterised by changes in their sensitivity to electrical stimulation of the proximal oesophagus following infusion of the distil oesophagus with acid. Method: The oesophagus of healthy volunteers (n=38) was intubated with a purpose built catheter to facilitate the irrigation of the distil oesophagus with acid (0.15M) and electrical stimulation of the proximal oesophagus. Additionally hospital based anxiety and depression questionnaires were completed prior to intubation. The pain threshold to moderate electrical stimulation (7 on a 0-10 VAS) was determined before and after oesophageal acidification. Blood samples were taken and DNA was extracted from volunteers characterised as sensitisers ( .6mA) or non sensitisers ( ,2mA) based on the change in post acid pain thresholds. Samples were screened for the pain protective GCH1 haplotype (Lotsch et al. 2007 Clin Chem 53(6): 1010-15) and the prevalence, pain threshold and depression/anxiety scores of volunteers were compared according to haplotype and pain phenotype. Data is expressed as mean ± sem and statistically analysed using one way ANOVA or Student's t-test as appropriate. Results: The prevalence of the pain protective haplotype (X) was O/O 66%, O/X 32%, X/X 3% overall. This prevalence was not significantly different between sensitisers vs non-sensitisers. Comparison of the pre-acid pain threshold between volunteers who possessed the pain protective allele (X) or did not (O) failed to reveal any significant difference (56.5 ± 7.2mA n=13 vs 51.8 ± 5.0mA n=25 p .0.05 respectively). However further analysis of the haplotype based on pain phenotype revealed that the baseline pain threshold in (X) sensitisers (76.6±5.6mA n=7) was significantly higher than other subgroups (58.6±6.4mA p=0.07 O sensitisers n=12; 33.0 ±4.2mA p ,0.01 X non-sensitisers n=6; 45.5±7.5mA p,0.05: O non-sensitisers n=13). Additionally depression scores were significantly lower in X sensitisers (7.1 ± 0.5) than the other groups (9.3 ± 0.4 p,0.01, 9.7 ± 0.7 p ,0.01, 8.8 ± 0.5 p ,0.05 respectively). No significant difference was found in anxiety scores. Conclusion: Preliminary data generated in the present study suggests that GCH-1 may play a role in visceral pain processing, particularly in subjects who sensitise to stimuli. This effect may be linked to psychosocial aspects, however further studies in larger cohorts are now warranted.
Introduction: Perception of the urge to defaecate is essential to normal evacuation. Anecdotal reports suggest an alteration in the ‘call to stool' in some constipated patients, particularly those with impaired rectal sensation (hyposensitivity) on diagnostic testing, but this has not been robustly tested. This study aimed to objectively assess pre-defaecatory sensations in patients with chronic constipation (CC) in comparison to asymptomatic volunteers (HV), and further to evaluate the role of rectal sensory status. Method: 100 patients with CC and 23 HV (all F) completed a standardized diary / questionnaire over a 5 day period, assessing presence (yes/no), quality (verbal descriptors), intensity (VAS) and location (area of anatomical diagram) of the urge to defaecate prior to attempted evacuation, as well as a sense of how effective evacuation was (VAS). Patients were stratified by rectal sensory status (to latex balloon distension) into those with hyposensitivity (RH: n=21), normal rectal sensation (NS: n=66), or hypersensitivity (n=13). All HV had normal rectal sensation. Results: Over the study period, mean defaecatory attempts /per subject were 4.3 in CC and 4.5 in HV (P= NS). On 13% of occasions, patients with CC visited the toilet in the absence of any urge to defaecate, compared to only 4% of visits by HV (P=0.0003). In both groups, urge to defaecate was described most commonly as "pressure/fullness" (CC=66% vs. HV=60%; P= NS), but patients used multiple other descriptive terms also: "aching" (23 vs. 10%); "cramping" (15 vs. 5%); "colic" (11 vs. 1%); "burning" (5 vs. 0%); "butterflies" (10 vs. 3%); "bloating" (35 vs. 10%); "nausea" (10 vs. 0%) [CC vs. HV, respectively: P ,0.05 for all). Generally, HV described a posterior sensation located in the anorectum (CC=31% vs. HV= 62%; P,0.0001), whereas CC were more like to describe a poorly localised sensation both anteriorly and posteriorly (CC = 41% vs. HV = 19%; P=0.0001). The area of viscerosomatic referral was greater in CC than HV (CC = 3% of diagram area [range 0-35%] vs. HV = 2% [0-8]; P,0.0001). Compared to constipated patients with NS, those with RH: (1) were less likely to describe the urge to defaecate as "pressure" or "fullness" (P=0.03 & P=0.001, respectively); (2) were more likely to report the urge as a sense of "nausea/sickness" (P= 0.01); (3) described a more diffuse area of sensation (P=0.001), predominately anteriorly (median RH = 2.6% vs. NS = 0.8%; P=0.003); (4) considered evacuation to be more effective (VAS in RH = 5.0 vs. NS = 2.8; P=0.004). Conclusions: This study confirms that patients with constipation have altered perception (location, quality and strength) of the urge to defaecate, with the greatest alteration seen in those with rectal hyposensitivity. Whether sensory dysfunction should be regarded a therapeutic target in constipated patients warrants consideration.
Tu2123 Spinal Cord Electrical Stimulation Improved Gastric Hypersensitivity in Rodent Models of Hyperalgesia Yan Sun, Geng-Qing Song, Jiande Chen
Tu2121 Mechanotransduction in Mouse Visceral Afferent Fibres Is Modulated by Nav1.9 James R. Hockley, Vincent Cibert-Goton, George Boundouki, John Wood, Charles H. Knowles, Mark D. Baker, Wendy Winchester, David C. Bulmer
Background: Spinal Cord Electrical Stimulation (SCS) has been used for treating chronic pain for many years. Visceral hypersensitivity (VH) is one of major pathogeneses of gastrointestinal diseases. VH has been established in a rodent model of gastric ulcer and a recent model of functional dyspepsia (FD). Aims: The aim of this study was designed to assess and compare the SCS effects on VH in two different rodent models, FD and gastric ulcer with gastric hyperalgesia. Methods: Experimental 1 (FD rats): 10 days old male Sprague-Dawley (SD) rats received 0.2 ml 0.1% iodoacetamide (IA) in 2% sucrose daily by oral gavages for 6 days. Rats were then allowed to grow normally to adult age (8-11 weeks) at which point electromyogram (EMG) and autonomic functions, and their responses to gastric distension (GD) and SCS were measured. Each rat served as its own control and underwent a number of randomized sessions on different days. SCS of different parameters (Pulse width: 0.08 2ms; frequency: 20 - 2000Hz; Amplitude: 90% motor threshold) were delivered at T9-T10. Autonomic functions were assessed by the spectral analysis of heart rate variability that was derived from the electrocardiogram signal. Experimental 2 was the same as Exp. 1 except that VH was established by injection of 10μl of 20% acetic acid into 10-12 sites in the submucosal layer of the stomach wall in 8-11 weeks old SD rats. Results: 1) In both models of VH, SCS with 0.2-0.5ms/50-100Hz resulted in substantial reduction of EMG by 36.8 48.8% (P,0.05 at 60 mmHg and 80 mmHg vs. the corresponding baseline); and SCS with 0.025ms-0.08ms/6000-20000Hz caused a mild reduction by 15.5 - 18.4% (P ,0.05 at 60 mmHg and 80 mmHg vs. the corresponding baseline); however, SCS with 0.2ms/20Hz, 1ms/ 100Hz, 2ms/100Hz, 3ms/100Hz showed little effects on EMG (P .0.05 vs. the corresponding baseline). 2) SCS with appropriate parameters (0.2ms/50Hz) improved GD-induced impaired sympathovagal balance (P,0.05) in both FD and gastric ulcer rats. 3) SCS with appropriate parameters (0.2ms/50Hz) reduced GD-induced increase of serum NE by 85.7% and 91.3% in FD and gastric ulcer rats, respectively (P ,0.05). Conclusions: SCS at T9-T10 with appropriate parameters reduces gastric hyperalgesia induced by gastric treatment of IA during neonatal stage or gastric treatment of acetic acid during adulthood; the inhibitory effect may be attributed to decreased sympathetic activity.
Mechanical stimulation of the bowel either by obstruction or distension is a principle cause of pain in functional and inflammatory bowel conditions, particularly in combination with visceral hypersensitivity. The voltage gated sodium channel subtype 1.9 (Nav1.9) has been implicated in the generation of mechanical hyperalgesia in models of inflammatory pain; however the role of Nav1.9 in visceral afferent sensitivity to mechanical distension in the gut has not been studied in great detail. Using Nav1.9 -/- mice we show that Nav1.9 has an important role in determining afferent activity to distension ex vivo. The distal colon and associated mesentery from C57BL6, Nav1.9 +/+ or Nav1.9 -/- mice were removed from mice euthanized by rising concentration of CO2, and placed in a recording chamber superfused with carbogenated Krebs buffer at 32-34°C supplemented with nifedipine (10μM), atropine (10μM) and indomethacin (3μM). The lumbar splanchnic nerve bundles were dissected free and nerve activity was recorded using a suction electrode. The tissue was luminally perfused with buffer (0.1mL/min) and the afferent response to ramp distensions (0-80mmHg or 0-145mmHg) determined. In separate preparations the afferent fibre response to repeated (10min interval) phasic 80mmHg distensions (60secs duration) were examined. Data is expressed as mean ± S.E.M. and statistically compared using a 2-way ANOVA or ttest, where appropriate. In Nav1.9 +/+ mice ramp distensions of 80mmHg evoked pressuredependent increases in colonic afferent firing producing a maximum response of 13.3 ± 3.7Hz (n=6) at 80mmHg which was greatly attenuated in recordings from Nav1.9 -/- mice (1.4 ± 0.6Hz n=6; P,0.05). By contrast no significant difference was seen in the maximum response to ramp distensions of 145mmHg between tissue from Nav1.9 +/+ mice (27.6 ± 3.1Hz n=5) compared with Nav1.9 -/- mice (25.5 ± 10.1Hz n=4). Similarly afferent responses to the first 80mmHg phasic distension were comparable in tissue from Nav1.9 +/+ and Nav1.9 -/- (n=4-5) mice. In Nav1.9 +/+ mice, further phasic distensions (2nd - 3rd), led to a decrease in magnitude of the afferent response, reaching a stable response during the 4th - 6th distensions (6th, 31.8 ± 6.9Hz). The decrease in afferent response to repeat distensions was more rapid in Nav1.9 -/- mice (6th; 6.0 ± 3.1Hz; P ,0.05) and failed to reach a stable response. Together these data demonstrate that Nav1.9 significantly contributes to colonic afferent mechanosensitivity dependent on the magnitude, frequency and dynamics of the stimulus.
Tu2124 P2X7 Receptor Modulate Primary Afferent Responses to Colorectal Distension in Rat Models With Visceral Pain Jing Wang, Elie D. Al-Chaer There is abundant evidence that microglia have a crucial role in the development and progression of various neurological disorders and response to persistent nociceptive stimulation. Extracellular adenosine 5'-triphosphate (ATP), one of the signaling molecules in the spinal cord plays an important role in the regulation of neuronal and glial functions in the nervous system through P2 purinoceptors. We previously reported that intrathecal injection of TNP-ATP - a P2X1-4 receptor antagonist, of PPADS - a P2X1-3,5 receptor antagonist,
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AGA Abstracts
AGA Abstracts
visceral analgesia observed with asimadoline in D-IBS patients with at least moderate pain. Supported by Tioga Pharmaceuticals and NHMRC Australia.
AGA Abstracts
nociceptive neurons expressed ASICs (ASIC1, ASIC2 and/or ASIC3, .40% nociceptive neurons of each subtype expressed all three genes). ASIC1 was expressed in 65-75% of the nodose, jugular and DRG nociceptive neurons. Both ASIC1 splice variants ASIC1a (75-80%) and ASIC1b (50-70%) were detected in the ASIC1-positive neurons. The distribution of ASIC1a and ASIC1b was similar between all nociceptive subtypes. ASIC2 was expressed in 50-70% of the nodose, jugular and DRG nociceptive neurons. The ASIC2 splice variant ASIC2b was detected in the majority of the nodose, jugular and DRG ASIC2-positive neurons. However, the splice variant ASIC2a was only detected in the nodose neurons ( .75% of ASIC2-positive nociceptive neurons), and was absent in the jugular and DRG neurons. ASIC3 was expressed in 70-75% of the nodose, jugular and DRG nociceptive neurons. In contrast to the guinea pig results, ASIC3 was not expressed in the mouse vagal and DRG esophageal nociceptive neurons. We conclude that the majority of the guinea pig esophageal nociceptive afferent neurons express ASIC1, ASIC2 and ASIC3 genes. While the ASIC1a, ASIC1b and ASIC2b splice variants are similarly expressed by the placodes-derived nodose and the neural crest-derived jugular and DRG nociceptive neurons, the ASIC2a subunit is only expressed in the nodose nociceptive subtype. Since the presence of ASIC2a subunit was found to substantially (≈10-fold) increase the threshold for acid in the multimeric ASIC channels, this ASIC2a expression pattern predicts that the nodose nociceptors will be less sensitive to acid via the ASIC mechanisms than their jugular and DRG counterparts. Our data also indicate an important difference in the ASIC expression in the esophageal nociceptive subtypes between the guinea pig and mouse. Supported by CEVYPET and ESF (ITMS: 26110230031).
and of Brilliant Blue G (BBG) - a P2X7 receptor antagonist significantly inhibited EMG responses to noxious colorectal distension (CRD) in rats with neonatal colon irritation (CI) or acute colon inflammation. In this study, we tested the effect of BBG on primary afferent responses to CRD in rat models of visceral pain. Experiments were conducted on adult male Sprague-Dawley rats pre-treated with neonatal CI, or which had acute colon inflammation secondary to adult treatment with TNBS. The spinal cord was exposed by laminectomy at the L5-S2 levels. The L6-S1 dorsal roots were dissected free from surrounding tissue. The dorsal rootlets were cut centrally, and their distal ends were carefully split into small filaments. Recording was done using a silver unipolar hook electrode placed on the distal stump of the cut dorsal rootlet filaments. Dorsal root potentials (DRP) were amplified and observed on a digital oscilloscope. Digitized signals were processed by an interface (CED 1401) connected to a PC to construct rate histograms. Spike 2 software was used to capture the original spikes as wavemark after subtracting the noise level. Responses to CRD were recorded before and 30 minutes after injection of BBG (25-75mg/kg i.v. Sigma). Results: 1. BBG (2575mg/kg i.v.) had no significant effects on DRP response to CRD in control rats (n=13). 2. In CI rats, BBG (50mg/kg i.v.) significantly decreased DRP response to noxious CRD by 40±10%, p,0.05 (60 mmHg) and 42.5±6%, p,0.05 (80mmHg) (n=5); BBG (75mg/kg i.v.) also significantly decreased DRP response to noxious CRD by 50±8%, p ,0.05 (60 mmHg) and 54.5±8%, p,0.05 (80mmHg) (n=3). Lower doses of BBG (25mg/kg i.v.) had no significant effect on DRP. 3. In TNBS-treated rats, BBG (50mg/kg i.v.) significantly decreased DRP response to noxious CRD by 45±8.4%, p ,0.05 (60 mmHg) and 44±7.2%, p ,0.05 (80mmHg) (n=4); BBG (75mg/kg i.v.) also significantly decreased DRP response to noxious CRD by 54±9%, p,0.01 (60 mmHg) and 47±8.5%, p ,0.05 (80mmHg) (n=4). Our results suggest that the P2X7 may play an important role in pain signaling, possibly through the activation of spinal cord microglia and may contribute to the generation of visceral hypersensitivity in rats with CI or with acute colon inflammation. (Supported by NIH / DK077733, DK081628 and DK081845)
Tu2127 Relationship Among Interleukin-6, Visceral Fat, and Colorectal Cancer Development Evaluated by Computed Tomography (CT) Colonography Yoshiki Kimura, Hiroshi Matsumoto, Ken Haruma
Tu2125
Background and Aims: Obesity, visceral fat is associated with increased risks for colorectal neoplasm, cancer (CRC) and adenomatous polyp (polyp). CT colonography is adopted as a new examination tool for colorectal cancer screening. The aim of this study was to evaluate the association between colorectal neoplasm and adiposity measurements assessed clinically and by CT. Methods: We prospectively evaluated 85 patients (mean age 61.2yrs, 50 men) referred to perform both colonoscopy and CT colonography. We devised three patient groups: no lesion (NL; n=46, men 27), polyp (polyp; n=25, men 13), and early CRC (CRC; n=14, men 10) groups. Visceral adiposity areas were determined at umbilical level by CT. Additionally, we also measured waist circumferences, serum lipid level and some plasma adipocytokines; adiponectin, leptin, TNF-α, IL-6 levels. Results: The mean triglyceride levels were higher in polyp group (147.7 mg/ml) than NL (98.4 ug/ml, p ,0.05). The mean uric acid levels were higher in polyp group (5.9 mg/ml) than NL (5.1 mg/ml, p ,0.05). The mean viscera adiposity area were higher in polyp group (159.6 cm2) than NL (114.3cm2, p,0.05). Moreover, mean serum IL-6 levels were lower in early CRC group (55.6 pg/ml ) than NL group (92 pg/ml, p ,0.05). Conclusion: Triglyceride, uric acid, viscera adiposity area may be intimately related to the primary incident of colorectal neoplasm. Moreover, IL-6 may be intimately related to the primary incident of colorectal cancer. We suggest further studies may be warranted to assess these adipocytokines as a novel target for colorectal neoplasm.
Refractory GERD Patients Display Increased Visceral Hypersensitivity for Thermal, Chemical and Mechanical Esophageal Stimulation Veerle Boecxstaens, Ans Pauwels, Kathleen Blondeau, Pantelis Oustamanolakis, Ege Altan, Guy E. Boeckxstaens, Jan F. Tack Background: In humans, pain is a multimodal experience composed of sensory, physiological and psychological aspects and esophageal sensitivity can be examined using different stimulation modalities. In gastro-esophageal reflux disease (GERD) patients with persistent symptoms in spite of PPIs, ongoing weakly acidic reflux is a well-established factor. Since reflux parameters in these patients are usually within the physiological number during PPI therapy, we speculate that visceral hypersensitivity might play a role in symptom perception. The aim of this study was to explore differences to multimodal sensory stimulation of the esophagus in healthy volunteers (HV) and GERD patients. Methods: 10 GERD patients (age 38y, 5m/5f) were compared to 10 HV (age35y, 4m/6f). Patients with typical GERD symptoms that persisted in spite of PPI b.i.d., were studied while on PPI. After an overnight fast, a multimodal stimulation probe was placed through the mouth with the balloon positioned 10cm proximal to the lower esophageal sphincter. Thermal (recirculating a saline solution through the balloon), mechanical (distensions of the balloon, with a maximal volume of 50cc), electrical and chemical (modified Bernstein test) stimulation of the esophagus were performed in all subjects in a semi-recumbent position. The sensory intensity was assessed by using a visual analogue scale (VAS) and stimulus intensities were increased with identification of first perception (VAS 1), pain threshold (VAS 5) up to the level of moderate pain (VAS 7), except for the electrical stimulation where only the pain threshold (VAS 5) was reached for safety reasons. All subjects filled out the Positive and Negative Affect Schedule (PAS and NAS) and State-Trait Anxiety Inventory (STAI) questionnaire before and after the stimulations. Results: Patients reached VAS 7 significantly earlier compared to HV for thermal stimulation (p=0.033), and were more likely than HVs to reach the VAS 7 during mechanical as well as chemical stimulation (p=0.017 and p=0.019 respectively, fig 1 and 2). No differences were observed between patients and HV in sensitivity to electrical stimulation. Patients had significantly higher NAS- and STAI-scores compared to HV prior to the test (p=0.0075 and p=0.0056 respectively), but none of these questionnaire scores were correlated with the sensitivity thresholds. Conclusions: In this study, we showed that GERD patients with persistent symptoms on PPI are hypersensitive to thermal, mechanical and chemical esophageal stimulation, compared to HV. These data confirm visceral hypersensitivity as an important candidate therapeutic target in PPI-refractory GERD.
Tu2128 Relationship Among Interleukin-6, Visceral Fat, and Colorectal Cancer Development Evaluated by Computed Tomography (CT) Colonography Yoshiki Kimura Background and Aims: Obesity, visceral fat is associated with increased risks for colorectal neoplasm, cancer (CRC) and adenomatous polyp (polyp). CT colonography is adopted as a new examination tool for colorectal cancer screening. The aim of this study was to evaluate the association between colorectal neoplasm and adiposity measurements assessed clinically and by CT. Methods: We prospectively evaluated 85 patients (mean age 61.2yrs, 50 men) referred to perform both colonoscopy and CT colonography. We devised three patient groups: no lesion (NL; n=46, men 27), polyp (polyp; n=25, men 13), and early CRC (CRC; n=14, men 10) groups. Visceral adiposity areas were determined at umbilical level by CT. Additionally, we also measured waist circumferences, serum lipid level and some plasma adipocytokines; adiponectin, leptin, TNF-α, IL-6 levels. Results: The mean triglyceride levels were higher in polyp group (147.7 mg/ml) than NL (98.4 ug/ml, p ,0.05). The mean uric acid levels were higher in polyp group (5.9 mg/ml) than NL (5.1 mg/ml, p ,0.05). The mean viscera adiposity area were higher in polyp group (159.6 cm2) than NL (114.3cm2, p,0.05). Moreover, mean serum IL-6 levels were lower in early CRC group (55.6 pg/ml ) than NL group (92 pg/ml, p ,0.05). Conclusion: Triglyceride, uric acid, viscera adiposity area may be intimately related to the primary incident of colorectal neoplasm. Moreover, IL-6 may be intimately related to the primary incident of colorectal cancer. We suggest further studies may be warranted to assess these adipocytokines as a novel target for colorectal neoplasm. Tu2129
Tu2126
Adipose Tissue Promotes the Growth and Invasion of Esophageal Squamous Cell Carcinoma and Inhibits It's Apoptosis Under Adipose Tissue-Cancer Cell Interaction In Vitro Atsushi Nakayama, Ryuichi Iwakiri, Shuji Toda, Kazuma Fujimoto
The Expression of the Acid Sensing Ion Channels mRNA in the Esophageal Afferent Nerves Svetlana Grobarcikova, Fei Ru, Jozef Hatok, Jan Kliment, Marian Kollarik The acid is a key noxious mediator in the esophagus, but the mechanisms of acid-induced activation in the esophageal afferent nerves are incompletely understood. We addressed the hypothesis that the esophageal nociceptive nerve subtypes express acid sensing ion channels (ASIC). Single cell RT-PCR was performed on the vagal nodose, vagal jugular and the spinal dorsal root ganglia (DRG) neurons retrogradely labeled from the esophagus in the guinea pig and mouse. Putative nociceptive neurons were identified by the TRPV1 expression. In the guinea pig nearly all (.90%) nodose (n=31), jugular (n=31) and DRG (n=23) esophageal
AGA Abstracts
Esophageal cancer develops within squamous epithelial layer, and progressively invades into submucosal to subadventitial layers. Adipose tissue exists in esophageal subadventitia. Thus, adipose tissue seems critical for the progression of esophageal cancer, but their interaction is unknown. Here we show the interaction, using electron microscopy, immunohistochemistry, Western blot, ELISA, real-time RT-PCR, and collagen gel invasion assay system, in which
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Tu2120 Quality, Intensity and Location of the Urge to Defaecate Differs Between Patients With Constipation and Healthy Subjects Rebecca Burgell, Michal M. Szczesniak, Priyanka Yerragorla, Emma V. Carrington, Sahar Mohammed, Peter J. Lunniss, Philip G. Dinning, S. Mark Scott
Introduction: GTP Cyclohydrolase-1 (GCH-1) is an enzyme responsible for the de novo synthesis of tetra borohydrate (BH4), a cofactor in monoamine and nitric oxide synthesis. Genetic studies have identified the presence of a pain protective GCH-1 haplotype which is associated with reduced BH4 production and lower pain scores in chronic pain patients or healthy volunteers following algogenic stimuli. The aim of this study was to examine influence of GCH-1 haplotype in healthy volunteers characterised by changes in their sensitivity to electrical stimulation of the proximal oesophagus following infusion of the distil oesophagus with acid. Method: The oesophagus of healthy volunteers (n=38) was intubated with a purpose built catheter to facilitate the irrigation of the distil oesophagus with acid (0.15M) and electrical stimulation of the proximal oesophagus. Additionally hospital based anxiety and depression questionnaires were completed prior to intubation. The pain threshold to moderate electrical stimulation (7 on a 0-10 VAS) was determined before and after oesophageal acidification. Blood samples were taken and DNA was extracted from volunteers characterised as sensitisers ( .6mA) or non sensitisers ( ,2mA) based on the change in post acid pain thresholds. Samples were screened for the pain protective GCH1 haplotype (Lotsch et al. 2007 Clin Chem 53(6): 1010-15) and the prevalence, pain threshold and depression/anxiety scores of volunteers were compared according to haplotype and pain phenotype. Data is expressed as mean ± sem and statistically analysed using one way ANOVA or Student's t-test as appropriate. Results: The prevalence of the pain protective haplotype (X) was O/O 66%, O/X 32%, X/X 3% overall. This prevalence was not significantly different between sensitisers vs non-sensitisers. Comparison of the pre-acid pain threshold between volunteers who possessed the pain protective allele (X) or did not (O) failed to reveal any significant difference (56.5 ± 7.2mA n=13 vs 51.8 ± 5.0mA n=25 p .0.05 respectively). However further analysis of the haplotype based on pain phenotype revealed that the baseline pain threshold in (X) sensitisers (76.6±5.6mA n=7) was significantly higher than other subgroups (58.6±6.4mA p=0.07 O sensitisers n=12; 33.0 ±4.2mA p ,0.01 X non-sensitisers n=6; 45.5±7.5mA p,0.05: O non-sensitisers n=13). Additionally depression scores were significantly lower in X sensitisers (7.1 ± 0.5) than the other groups (9.3 ± 0.4 p,0.01, 9.7 ± 0.7 p ,0.01, 8.8 ± 0.5 p ,0.05 respectively). No significant difference was found in anxiety scores. Conclusion: Preliminary data generated in the present study suggests that GCH-1 may play a role in visceral pain processing, particularly in subjects who sensitise to stimuli. This effect may be linked to psychosocial aspects, however further studies in larger cohorts are now warranted.
Introduction: Perception of the urge to defaecate is essential to normal evacuation. Anecdotal reports suggest an alteration in the ‘call to stool' in some constipated patients, particularly those with impaired rectal sensation (hyposensitivity) on diagnostic testing, but this has not been robustly tested. This study aimed to objectively assess pre-defaecatory sensations in patients with chronic constipation (CC) in comparison to asymptomatic volunteers (HV), and further to evaluate the role of rectal sensory status. Method: 100 patients with CC and 23 HV (all F) completed a standardized diary / questionnaire over a 5 day period, assessing presence (yes/no), quality (verbal descriptors), intensity (VAS) and location (area of anatomical diagram) of the urge to defaecate prior to attempted evacuation, as well as a sense of how effective evacuation was (VAS). Patients were stratified by rectal sensory status (to latex balloon distension) into those with hyposensitivity (RH: n=21), normal rectal sensation (NS: n=66), or hypersensitivity (n=13). All HV had normal rectal sensation. Results: Over the study period, mean defaecatory attempts /per subject were 4.3 in CC and 4.5 in HV (P= NS). On 13% of occasions, patients with CC visited the toilet in the absence of any urge to defaecate, compared to only 4% of visits by HV (P=0.0003). In both groups, urge to defaecate was described most commonly as "pressure/fullness" (CC=66% vs. HV=60%; P= NS), but patients used multiple other descriptive terms also: "aching" (23 vs. 10%); "cramping" (15 vs. 5%); "colic" (11 vs. 1%); "burning" (5 vs. 0%); "butterflies" (10 vs. 3%); "bloating" (35 vs. 10%); "nausea" (10 vs. 0%) [CC vs. HV, respectively: P ,0.05 for all). Generally, HV described a posterior sensation located in the anorectum (CC=31% vs. HV= 62%; P,0.0001), whereas CC were more like to describe a poorly localised sensation both anteriorly and posteriorly (CC = 41% vs. HV = 19%; P=0.0001). The area of viscerosomatic referral was greater in CC than HV (CC = 3% of diagram area [range 0-35%] vs. HV = 2% [0-8]; P,0.0001). Compared to constipated patients with NS, those with RH: (1) were less likely to describe the urge to defaecate as "pressure" or "fullness" (P=0.03 & P=0.001, respectively); (2) were more likely to report the urge as a sense of "nausea/sickness" (P= 0.01); (3) described a more diffuse area of sensation (P=0.001), predominately anteriorly (median RH = 2.6% vs. NS = 0.8%; P=0.003); (4) considered evacuation to be more effective (VAS in RH = 5.0 vs. NS = 2.8; P=0.004). Conclusions: This study confirms that patients with constipation have altered perception (location, quality and strength) of the urge to defaecate, with the greatest alteration seen in those with rectal hyposensitivity. Whether sensory dysfunction should be regarded a therapeutic target in constipated patients warrants consideration.
Tu2123 Spinal Cord Electrical Stimulation Improved Gastric Hypersensitivity in Rodent Models of Hyperalgesia Yan Sun, Geng-Qing Song, Jiande Chen
Tu2121 Mechanotransduction in Mouse Visceral Afferent Fibres Is Modulated by Nav1.9 James R. Hockley, Vincent Cibert-Goton, George Boundouki, John Wood, Charles H. Knowles, Mark D. Baker, Wendy Winchester, David C. Bulmer
Background: Spinal Cord Electrical Stimulation (SCS) has been used for treating chronic pain for many years. Visceral hypersensitivity (VH) is one of major pathogeneses of gastrointestinal diseases. VH has been established in a rodent model of gastric ulcer and a recent model of functional dyspepsia (FD). Aims: The aim of this study was designed to assess and compare the SCS effects on VH in two different rodent models, FD and gastric ulcer with gastric hyperalgesia. Methods: Experimental 1 (FD rats): 10 days old male Sprague-Dawley (SD) rats received 0.2 ml 0.1% iodoacetamide (IA) in 2% sucrose daily by oral gavages for 6 days. Rats were then allowed to grow normally to adult age (8-11 weeks) at which point electromyogram (EMG) and autonomic functions, and their responses to gastric distension (GD) and SCS were measured. Each rat served as its own control and underwent a number of randomized sessions on different days. SCS of different parameters (Pulse width: 0.08 2ms; frequency: 20 - 2000Hz; Amplitude: 90% motor threshold) were delivered at T9-T10. Autonomic functions were assessed by the spectral analysis of heart rate variability that was derived from the electrocardiogram signal. Experimental 2 was the same as Exp. 1 except that VH was established by injection of 10μl of 20% acetic acid into 10-12 sites in the submucosal layer of the stomach wall in 8-11 weeks old SD rats. Results: 1) In both models of VH, SCS with 0.2-0.5ms/50-100Hz resulted in substantial reduction of EMG by 36.8 48.8% (P,0.05 at 60 mmHg and 80 mmHg vs. the corresponding baseline); and SCS with 0.025ms-0.08ms/6000-20000Hz caused a mild reduction by 15.5 - 18.4% (P ,0.05 at 60 mmHg and 80 mmHg vs. the corresponding baseline); however, SCS with 0.2ms/20Hz, 1ms/ 100Hz, 2ms/100Hz, 3ms/100Hz showed little effects on EMG (P .0.05 vs. the corresponding baseline). 2) SCS with appropriate parameters (0.2ms/50Hz) improved GD-induced impaired sympathovagal balance (P,0.05) in both FD and gastric ulcer rats. 3) SCS with appropriate parameters (0.2ms/50Hz) reduced GD-induced increase of serum NE by 85.7% and 91.3% in FD and gastric ulcer rats, respectively (P ,0.05). Conclusions: SCS at T9-T10 with appropriate parameters reduces gastric hyperalgesia induced by gastric treatment of IA during neonatal stage or gastric treatment of acetic acid during adulthood; the inhibitory effect may be attributed to decreased sympathetic activity.
Mechanical stimulation of the bowel either by obstruction or distension is a principle cause of pain in functional and inflammatory bowel conditions, particularly in combination with visceral hypersensitivity. The voltage gated sodium channel subtype 1.9 (Nav1.9) has been implicated in the generation of mechanical hyperalgesia in models of inflammatory pain; however the role of Nav1.9 in visceral afferent sensitivity to mechanical distension in the gut has not been studied in great detail. Using Nav1.9 -/- mice we show that Nav1.9 has an important role in determining afferent activity to distension ex vivo. The distal colon and associated mesentery from C57BL6, Nav1.9 +/+ or Nav1.9 -/- mice were removed from mice euthanized by rising concentration of CO2, and placed in a recording chamber superfused with carbogenated Krebs buffer at 32-34°C supplemented with nifedipine (10μM), atropine (10μM) and indomethacin (3μM). The lumbar splanchnic nerve bundles were dissected free and nerve activity was recorded using a suction electrode. The tissue was luminally perfused with buffer (0.1mL/min) and the afferent response to ramp distensions (0-80mmHg or 0-145mmHg) determined. In separate preparations the afferent fibre response to repeated (10min interval) phasic 80mmHg distensions (60secs duration) were examined. Data is expressed as mean ± S.E.M. and statistically compared using a 2-way ANOVA or ttest, where appropriate. In Nav1.9 +/+ mice ramp distensions of 80mmHg evoked pressuredependent increases in colonic afferent firing producing a maximum response of 13.3 ± 3.7Hz (n=6) at 80mmHg which was greatly attenuated in recordings from Nav1.9 -/- mice (1.4 ± 0.6Hz n=6; P,0.05). By contrast no significant difference was seen in the maximum response to ramp distensions of 145mmHg between tissue from Nav1.9 +/+ mice (27.6 ± 3.1Hz n=5) compared with Nav1.9 -/- mice (25.5 ± 10.1Hz n=4). Similarly afferent responses to the first 80mmHg phasic distension were comparable in tissue from Nav1.9 +/+ and Nav1.9 -/- (n=4-5) mice. In Nav1.9 +/+ mice, further phasic distensions (2nd - 3rd), led to a decrease in magnitude of the afferent response, reaching a stable response during the 4th - 6th distensions (6th, 31.8 ± 6.9Hz). The decrease in afferent response to repeat distensions was more rapid in Nav1.9 -/- mice (6th; 6.0 ± 3.1Hz; P ,0.05) and failed to reach a stable response. Together these data demonstrate that Nav1.9 significantly contributes to colonic afferent mechanosensitivity dependent on the magnitude, frequency and dynamics of the stimulus.
Tu2124 P2X7 Receptor Modulate Primary Afferent Responses to Colorectal Distension in Rat Models With Visceral Pain Jing Wang, Elie D. Al-Chaer There is abundant evidence that microglia have a crucial role in the development and progression of various neurological disorders and response to persistent nociceptive stimulation. Extracellular adenosine 5'-triphosphate (ATP), one of the signaling molecules in the spinal cord plays an important role in the regulation of neuronal and glial functions in the nervous system through P2 purinoceptors. We previously reported that intrathecal injection of TNP-ATP - a P2X1-4 receptor antagonist, of PPADS - a P2X1-3,5 receptor antagonist,
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AGA Abstracts
AGA Abstracts
visceral analgesia observed with asimadoline in D-IBS patients with at least moderate pain. Supported by Tioga Pharmaceuticals and NHMRC Australia.
AGA Abstracts
nociceptive neurons expressed ASICs (ASIC1, ASIC2 and/or ASIC3, .40% nociceptive neurons of each subtype expressed all three genes). ASIC1 was expressed in 65-75% of the nodose, jugular and DRG nociceptive neurons. Both ASIC1 splice variants ASIC1a (75-80%) and ASIC1b (50-70%) were detected in the ASIC1-positive neurons. The distribution of ASIC1a and ASIC1b was similar between all nociceptive subtypes. ASIC2 was expressed in 50-70% of the nodose, jugular and DRG nociceptive neurons. The ASIC2 splice variant ASIC2b was detected in the majority of the nodose, jugular and DRG ASIC2-positive neurons. However, the splice variant ASIC2a was only detected in the nodose neurons ( .75% of ASIC2-positive nociceptive neurons), and was absent in the jugular and DRG neurons. ASIC3 was expressed in 70-75% of the nodose, jugular and DRG nociceptive neurons. In contrast to the guinea pig results, ASIC3 was not expressed in the mouse vagal and DRG esophageal nociceptive neurons. We conclude that the majority of the guinea pig esophageal nociceptive afferent neurons express ASIC1, ASIC2 and ASIC3 genes. While the ASIC1a, ASIC1b and ASIC2b splice variants are similarly expressed by the placodes-derived nodose and the neural crest-derived jugular and DRG nociceptive neurons, the ASIC2a subunit is only expressed in the nodose nociceptive subtype. Since the presence of ASIC2a subunit was found to substantially (≈10-fold) increase the threshold for acid in the multimeric ASIC channels, this ASIC2a expression pattern predicts that the nodose nociceptors will be less sensitive to acid via the ASIC mechanisms than their jugular and DRG counterparts. Our data also indicate an important difference in the ASIC expression in the esophageal nociceptive subtypes between the guinea pig and mouse. Supported by CEVYPET and ESF (ITMS: 26110230031).
and of Brilliant Blue G (BBG) - a P2X7 receptor antagonist significantly inhibited EMG responses to noxious colorectal distension (CRD) in rats with neonatal colon irritation (CI) or acute colon inflammation. In this study, we tested the effect of BBG on primary afferent responses to CRD in rat models of visceral pain. Experiments were conducted on adult male Sprague-Dawley rats pre-treated with neonatal CI, or which had acute colon inflammation secondary to adult treatment with TNBS. The spinal cord was exposed by laminectomy at the L5-S2 levels. The L6-S1 dorsal roots were dissected free from surrounding tissue. The dorsal rootlets were cut centrally, and their distal ends were carefully split into small filaments. Recording was done using a silver unipolar hook electrode placed on the distal stump of the cut dorsal rootlet filaments. Dorsal root potentials (DRP) were amplified and observed on a digital oscilloscope. Digitized signals were processed by an interface (CED 1401) connected to a PC to construct rate histograms. Spike 2 software was used to capture the original spikes as wavemark after subtracting the noise level. Responses to CRD were recorded before and 30 minutes after injection of BBG (25-75mg/kg i.v. Sigma). Results: 1. BBG (2575mg/kg i.v.) had no significant effects on DRP response to CRD in control rats (n=13). 2. In CI rats, BBG (50mg/kg i.v.) significantly decreased DRP response to noxious CRD by 40±10%, p,0.05 (60 mmHg) and 42.5±6%, p,0.05 (80mmHg) (n=5); BBG (75mg/kg i.v.) also significantly decreased DRP response to noxious CRD by 50±8%, p ,0.05 (60 mmHg) and 54.5±8%, p,0.05 (80mmHg) (n=3). Lower doses of BBG (25mg/kg i.v.) had no significant effect on DRP. 3. In TNBS-treated rats, BBG (50mg/kg i.v.) significantly decreased DRP response to noxious CRD by 45±8.4%, p ,0.05 (60 mmHg) and 44±7.2%, p ,0.05 (80mmHg) (n=4); BBG (75mg/kg i.v.) also significantly decreased DRP response to noxious CRD by 54±9%, p,0.01 (60 mmHg) and 47±8.5%, p ,0.05 (80mmHg) (n=4). Our results suggest that the P2X7 may play an important role in pain signaling, possibly through the activation of spinal cord microglia and may contribute to the generation of visceral hypersensitivity in rats with CI or with acute colon inflammation. (Supported by NIH / DK077733, DK081628 and DK081845)
Tu2127 Relationship Among Interleukin-6, Visceral Fat, and Colorectal Cancer Development Evaluated by Computed Tomography (CT) Colonography Yoshiki Kimura, Hiroshi Matsumoto, Ken Haruma
Tu2125
Background and Aims: Obesity, visceral fat is associated with increased risks for colorectal neoplasm, cancer (CRC) and adenomatous polyp (polyp). CT colonography is adopted as a new examination tool for colorectal cancer screening. The aim of this study was to evaluate the association between colorectal neoplasm and adiposity measurements assessed clinically and by CT. Methods: We prospectively evaluated 85 patients (mean age 61.2yrs, 50 men) referred to perform both colonoscopy and CT colonography. We devised three patient groups: no lesion (NL; n=46, men 27), polyp (polyp; n=25, men 13), and early CRC (CRC; n=14, men 10) groups. Visceral adiposity areas were determined at umbilical level by CT. Additionally, we also measured waist circumferences, serum lipid level and some plasma adipocytokines; adiponectin, leptin, TNF-α, IL-6 levels. Results: The mean triglyceride levels were higher in polyp group (147.7 mg/ml) than NL (98.4 ug/ml, p ,0.05). The mean uric acid levels were higher in polyp group (5.9 mg/ml) than NL (5.1 mg/ml, p ,0.05). The mean viscera adiposity area were higher in polyp group (159.6 cm2) than NL (114.3cm2, p,0.05). Moreover, mean serum IL-6 levels were lower in early CRC group (55.6 pg/ml ) than NL group (92 pg/ml, p ,0.05). Conclusion: Triglyceride, uric acid, viscera adiposity area may be intimately related to the primary incident of colorectal neoplasm. Moreover, IL-6 may be intimately related to the primary incident of colorectal cancer. We suggest further studies may be warranted to assess these adipocytokines as a novel target for colorectal neoplasm.
Refractory GERD Patients Display Increased Visceral Hypersensitivity for Thermal, Chemical and Mechanical Esophageal Stimulation Veerle Boecxstaens, Ans Pauwels, Kathleen Blondeau, Pantelis Oustamanolakis, Ege Altan, Guy E. Boeckxstaens, Jan F. Tack Background: In humans, pain is a multimodal experience composed of sensory, physiological and psychological aspects and esophageal sensitivity can be examined using different stimulation modalities. In gastro-esophageal reflux disease (GERD) patients with persistent symptoms in spite of PPIs, ongoing weakly acidic reflux is a well-established factor. Since reflux parameters in these patients are usually within the physiological number during PPI therapy, we speculate that visceral hypersensitivity might play a role in symptom perception. The aim of this study was to explore differences to multimodal sensory stimulation of the esophagus in healthy volunteers (HV) and GERD patients. Methods: 10 GERD patients (age 38y, 5m/5f) were compared to 10 HV (age35y, 4m/6f). Patients with typical GERD symptoms that persisted in spite of PPI b.i.d., were studied while on PPI. After an overnight fast, a multimodal stimulation probe was placed through the mouth with the balloon positioned 10cm proximal to the lower esophageal sphincter. Thermal (recirculating a saline solution through the balloon), mechanical (distensions of the balloon, with a maximal volume of 50cc), electrical and chemical (modified Bernstein test) stimulation of the esophagus were performed in all subjects in a semi-recumbent position. The sensory intensity was assessed by using a visual analogue scale (VAS) and stimulus intensities were increased with identification of first perception (VAS 1), pain threshold (VAS 5) up to the level of moderate pain (VAS 7), except for the electrical stimulation where only the pain threshold (VAS 5) was reached for safety reasons. All subjects filled out the Positive and Negative Affect Schedule (PAS and NAS) and State-Trait Anxiety Inventory (STAI) questionnaire before and after the stimulations. Results: Patients reached VAS 7 significantly earlier compared to HV for thermal stimulation (p=0.033), and were more likely than HVs to reach the VAS 7 during mechanical as well as chemical stimulation (p=0.017 and p=0.019 respectively, fig 1 and 2). No differences were observed between patients and HV in sensitivity to electrical stimulation. Patients had significantly higher NAS- and STAI-scores compared to HV prior to the test (p=0.0075 and p=0.0056 respectively), but none of these questionnaire scores were correlated with the sensitivity thresholds. Conclusions: In this study, we showed that GERD patients with persistent symptoms on PPI are hypersensitive to thermal, mechanical and chemical esophageal stimulation, compared to HV. These data confirm visceral hypersensitivity as an important candidate therapeutic target in PPI-refractory GERD.
Tu2128 Relationship Among Interleukin-6, Visceral Fat, and Colorectal Cancer Development Evaluated by Computed Tomography (CT) Colonography Yoshiki Kimura Background and Aims: Obesity, visceral fat is associated with increased risks for colorectal neoplasm, cancer (CRC) and adenomatous polyp (polyp). CT colonography is adopted as a new examination tool for colorectal cancer screening. The aim of this study was to evaluate the association between colorectal neoplasm and adiposity measurements assessed clinically and by CT. Methods: We prospectively evaluated 85 patients (mean age 61.2yrs, 50 men) referred to perform both colonoscopy and CT colonography. We devised three patient groups: no lesion (NL; n=46, men 27), polyp (polyp; n=25, men 13), and early CRC (CRC; n=14, men 10) groups. Visceral adiposity areas were determined at umbilical level by CT. Additionally, we also measured waist circumferences, serum lipid level and some plasma adipocytokines; adiponectin, leptin, TNF-α, IL-6 levels. Results: The mean triglyceride levels were higher in polyp group (147.7 mg/ml) than NL (98.4 ug/ml, p ,0.05). The mean uric acid levels were higher in polyp group (5.9 mg/ml) than NL (5.1 mg/ml, p ,0.05). The mean viscera adiposity area were higher in polyp group (159.6 cm2) than NL (114.3cm2, p,0.05). Moreover, mean serum IL-6 levels were lower in early CRC group (55.6 pg/ml ) than NL group (92 pg/ml, p ,0.05). Conclusion: Triglyceride, uric acid, viscera adiposity area may be intimately related to the primary incident of colorectal neoplasm. Moreover, IL-6 may be intimately related to the primary incident of colorectal cancer. We suggest further studies may be warranted to assess these adipocytokines as a novel target for colorectal neoplasm. Tu2129
Tu2126
Adipose Tissue Promotes the Growth and Invasion of Esophageal Squamous Cell Carcinoma and Inhibits It's Apoptosis Under Adipose Tissue-Cancer Cell Interaction In Vitro Atsushi Nakayama, Ryuichi Iwakiri, Shuji Toda, Kazuma Fujimoto
The Expression of the Acid Sensing Ion Channels mRNA in the Esophageal Afferent Nerves Svetlana Grobarcikova, Fei Ru, Jozef Hatok, Jan Kliment, Marian Kollarik The acid is a key noxious mediator in the esophagus, but the mechanisms of acid-induced activation in the esophageal afferent nerves are incompletely understood. We addressed the hypothesis that the esophageal nociceptive nerve subtypes express acid sensing ion channels (ASIC). Single cell RT-PCR was performed on the vagal nodose, vagal jugular and the spinal dorsal root ganglia (DRG) neurons retrogradely labeled from the esophagus in the guinea pig and mouse. Putative nociceptive neurons were identified by the TRPV1 expression. In the guinea pig nearly all (.90%) nodose (n=31), jugular (n=31) and DRG (n=23) esophageal
AGA Abstracts
Esophageal cancer develops within squamous epithelial layer, and progressively invades into submucosal to subadventitial layers. Adipose tissue exists in esophageal subadventitia. Thus, adipose tissue seems critical for the progression of esophageal cancer, but their interaction is unknown. Here we show the interaction, using electron microscopy, immunohistochemistry, Western blot, ELISA, real-time RT-PCR, and collagen gel invasion assay system, in which
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