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Localization of cannabinoid (CB1) receptor in hindbrain and nodose ganglion of ferrets
612 Localization of Cannabinoid (CB1) Receptor in Hindbrain and Nndose Ganglion of Ferrets Elita R. Partosoedarso,Roisin T. Scullion, Thomas P. Abrahams, PamelaJ. Hornby, LSU Health Science Ctr, New Orleans, LA BACKGROUND. Delta9- tetrahydrocannabinol (Ag-THC), an active constituent of marijuana, has marked effects on gastrointestinal motility and is sometimes used as an anti-emetic in humans. In rats, systemic administrationof L~9-THCinhibits gastric contractility and this effect is abolished by vagotomy or a cannabinoidl receptor (CB1R) antagonist (Eur.J.Pharmecol. 371:187,1999). Thesedata suggestthat~9-THC acts on CBt R associatedwith vagal pathways that control gastric motor function. Therefore, we used immunocytochemistry and a wellcharacterized CB1R antibody (Neuroscience 83:393,1998) in ferret hindbrain and nodose ganglion to assessthe site of action of CB1R for this effect. METHODS.Sections of hindbraln and nodose ganglion were incubated with antiserum to the rat construct of CBIR (1:10001:4000), which was followed by tyramide amplification (TSA-flourescein Idt;, NEN, Boston, MA) of the signal. RESULTS.CB1R immunostaining was visualized in neurons in the area postrema and their processes. Intense labeling was visualized in some cells in the nucleus tractus solitarius (NTS), as well as a diffuse punetate labeling throughout the NTS and the dorsal motor nucleus of the vagus (DMN). CB1R was never cctocalizedwith cholera toxin in DMN neurons after the retrograde label was injected into the fondus smooth muscle. In addition, vagotomy performed 7 days previously did not alter CB1R staining in the lOs/lateral dorsal vagal complex (whereas, a loss of staining would be expected if the receptor was expressed by vagal motor neurons). Therefore, CB1R is not expressedin vsgal preganglionic neurons innervating the fundus. Moderate CB1R staining was observed in cells within the nodose ganglion; however,7 days after no(lose ganglionectomythe intensity of CB1R staining in the ipsilateral NTS was not noticeably different from control side. CONCLUSIONS.These data suggest that the majority of CB1R staining in the NTS does not arise from primary afferents. In addition, since L~9-THCapplied to the dorsal surface of the medulla mimics the effects of systemic L~9-THC(Eur.J.Pharmacol.371:187,1999), we conclude that peripherally administered Ag-THCcould act on CB1Rsin the area postremaand NTS to alter gastric motor function. Finally, L~9-THCis unlikely to directly affect motor output from preganglionic vagal neurons innervating the stomach. Supported by DK43714 (PJH) and ADHF (TPA).
613 Effects Of Stimulating Protease-Activated Rncoptors On Myatodc ~ in Guinea-Pig Small intestine Chuanyun Gag, Hong-Zhen Hu, Sumei L/u, Na Gag, Yun X/a, Jackie D. Wood, Ohio State Univ, Columbus, OH Background:Protease-activatedreceptors (PARs) are a family of G-protein coupled receptors that are cleaved and activated by certain proteases. Four members have been cloned and designated: PAR-l, 2, 3, 4. Among them, PAR-1 and PAR-2 have been localized in tissues of the gastrointestinal tract by immunohistochemistry and their roles in intestinal disorders assoclatad with inflammation have been implied. Nevertheless,the exact mechanism of an'don has not been fully clarified. This study tested the hypothesis that proteases modulate the gastrointestinalsystemthrough actions on the enteric nervous system. Methods: Conventional intracellular microelectrode and neuron tracer injection techniques were used to study the action of thrombin, trypsin, tryptase, PAR-1 tethered ligand analogue SFLLRL, and PAR-2 tethered ligand analogue SRLIGL on myenteric neurons of the guinea-pig small intestine. Results: Bath application of thrombin (O.l-lOunits/lOml) or trypsin (1-100 nM) evoked s tetrodoxin-resistant,concentration-dependentdepolarizingresponsesin meet of the myenteric neurons examined (77% in 56 S-type neurons, and 75% of 89 AH-type neurons). The depolarizationwas accompaniedby decreasedinput resistanceand associatedwith increased excitability reflected by repetitive spike discharge during depolarizing current pulses and anode-breakexcitation at the offset of hyperpolarizingcurrent pulses. Application of SFLLRI., the PAR-1 agonist, mimicked the effect of thrombin and trypsin with reduced potency of 0.11 p.M. In comparison,tryptase (0.1-10 nM) or PAR-2anon/st, SLIGRL, only produced a weak depolarizationof 2-4mV. The cyclooxygenaseinhibitor piroxicam (60 mM) did not affect the depolarizing responses produced by thrombin, trypsin or SFLLRL. Conclusion: The results suggest that the excitatory actions of proteasesare mediated primarily by PAR-t. PAR-2 may also contribute to the excitatory action of proteases,however,to a lesser extent. Arachidonic acid metabolism is not involved in the protease-evokedactivation of enteric neurons. (Supported by NIH R01 DK37238
614 is Psychiatric Illness, Personality or Both a Predictor of IBS in Young Adults? Results from a Birth Cohort Study in New Zealand. Stuart C. Howell, Univ of Sydney, Sydney Australia; Richie Pouifon, Univ of Otugo, Ounedin New Zealand; Nicholas J. Talley, Univ of Sydney, Sydney Australia Background:The association of IBS with psychiatric disorder remains uncertain. Relationships have been reported in some cross-sechonal studies, however, these have failed to establish whether psychiatric disorder precedes a diagnosis of IBS or is a consequence of if. Few studies haveexaminedwhether psychiatric disorder is more important than personalityfactors in the etiology of IBS. Method: The data were collected from a 1972 birth cohort (Dunedin, NZ), which has beenfollowed through the first 26 years of life: 96% of the living cohort was assessed at the most recent follow-up (at age 26). IBS was diagnosed according to the Manning criteria, using symptom data collected at age 26. DSM-IIIR diagnostic information was collected at ages 18 and 21 years; a DSM diagnosis at both time points was used to define chronic psychiatric disorder (any), chronic depressive illness and chronic anxiety disorder. Personality was assessed at age 18 years using the Multidimensional Personality Questionnaire (MPQ). Results: Datawas availablefor 869 subjects (49.6% female). Of these, 145 (18.7%) met the Manning criteria for IBS. This was significantly more common among females compared to males (19.5% vs 13.9%; p=O.03), but not among individuals with a history of psychiatric disorder. The lack of effect for psychiatric disorder was evident for any
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disorder (p=0.72), and for depressive illness (p=0.10) and anxiety disorder (p~0.53). Of the ten MPQ scales considered, only one emerged as a significant predictor of a diagnosis of IBS: mean scores on the Social Potency scale were significantly higher among subjects with IBS when compared to normal controls (Mean: 42.2 vs 36.6; p =0.02). The difference remained significant after adjustment for subject gender (p=O.01). Conclusions: In young adults, IBS is relatedto personality,but not to a history of psychiatricdisorder. When compared to healthy controls, IBS patients were significantly more likely to report a socially 'forceful' or dominant personality type.
615 Ps~intric Oiagnmms in GeatroenteralOy: Validation of a SeHoReporl Instrument (PRIME-MO Patient Health Qumdioneaira), Epidomiolo0y and Recognition, Philippe Persoons, Koen Luyckx, Benjamin Fischler, KatholiekeUniv Leuven, Dept Lialsonpcyohiatne, Leuven Belgium PURPOSE: In Gastroenterology(GE) no serf-report instrument to assess Anxiety and Mood Disorders (AMD) exists. We validated the AMD modules of the PRIME-MD Patient Health Questionnaire (PHO; Spitzer et al., JAMA 1999), the first categorial, self-report measure of psychialdc disorders, in GEpatientsand assessedthe epidemiologyand recognitionby medical residents of AMD in GE inpatients. METHODS: 639 consecutive GE in- and outpatients completed the PHQ. 184 patients were interviewed by a mental health professional with the Mini International Neoropsychlatric Interview 5.O.O to assess criterion validity. Construct validity was assessedwith the SF36 (Ouality of Life), Hospital Anxiety and Depressionscale (HAl)S), Beck Depression Inventory 7-items (BDI-PC), health care use and disability days. A questionnaireon the presenceof AMD in 175 inpatients was completed by medical residents, blind for the PHQ. Inpatients (n=475) were divided into 3 groups: Functional Intestinal Disorder (FID;n=87), Inflammatory Bowel Disease (IBO;n=lOl), Other Organic Disorder (OOD;n=287). RESULTS: Sensitivity and specificity were respectively 80.0% and 98.1% (K = 0.81) for Major DepressiveDisorder (MDD), 83.0% and 91.5% (K = 0.74) for Any Anxiety and DepressiveOisorder (AADO), 77.1% and 91.9% (K= 0.69) for Any DepressiveOisorder (ADD), 70.8% and 94.2% (K = 0.65) for Any Anxiety Disorder (AAD). Construct validity was satisfying and operating characteristics were similar or better than for HADS and BDI-PC. Prevalenceof AADD,ADD, MDD, Other DepressiveDisorder (ODD), AAD, Panic Disorder and Other Anxiety Disorder in inpatients was respectively36.0%, 31.4%, 17.9%, 13.5%, 14.9%, 6.8% and 9.6%. In lid there were significantly higher prevalences than in IBO and OOD (except for ODD) and in IBO there were significantly higher prevalencesfor AAOOand MOD than in OOD. Recognition rates by medical residents of AADO, ADO, MOD, ODD and AAD were respectively 27.0%, 22.7%, 10.8%, 13.8% and 20.7%. Recognition for AADD was significantly better in RO than in ISO and GOD. CONCLUSION:AMD are highly prevalent in GE inpatlents, especially in FID and IBD. AMO are poorly recognized by medical residents, especially in OOD and IBD. The PHG appears to be a diagnostic instrument for MDD and a good screening instrument for ADO and AAD in GE patients. The systematic use of the PHQ couM improve the recognition of AMD on GE wards.
616 Ookavioral aM Pl~ological Sleep Characteristics in IBS Symptom Subgroups S/god Elsonbruch, Dept of Medical Psychology, Univ Clin of Essen, Essen Germany; Jennifer J. Thompson, Michael J. Harnish, William C. Orr, Lynn Institute for Heaithcare Research, Ok/shores C/t/, OK BACKGROUND:Although subjective sleep complaints are well-documented in patients with IBS, the presence of objective sleep abnormalities remains controversial. It also remains unclear whether sleep dysfunction is relatedto patients gastrointestinal (GI) symptoms or to psychological factors. The goal of this study was to investigate sleep characteristics in IBS symtpom subgroups. METHODS:31 female/BS pts were stratified into 15 pts with only bowel symptoms (IBSonly), and 16 pts with both lower and upperdyspeptic-likesymptoms (IBS+ D). 21 healthy females served as controls. For 4 consecutive days subjective sleep quality, insomnia symptoms, alertness, state anxiety, stress levels, and daytime and night time GI symptoms were assessed. Saliva samples were collected for curt/sol analysis. On night 4, subjects underwent polysomnographic(PSG) moniforJngfor an objectiveassessmentof sleep quality including arousals and respiratory parameters. RESULTS:Comparedto both IBS-only and controls, IBS + D reported greater difficulties falling asleep, and showed a greater global Pittsburgh Sleep Quality Index (PSQI) score indicating greater overall sleep problems (all p<.05). A greater percentageof IBS + Opts reported night time GI symptoms (73.5 vs. 38.25 %, p<.01). Compared to both controls and IBS-only, IBS+D reported greater depression and morning anxiety across the 4 study (lays (all p<.05). Both patient groups felt less rested and showed decreasedlevels of daytime alertness on all 4 days compared to controls (all p<.01). Both patient groups showed normal circadian changes in salivary curt/sol levels (evening: 4.2 _+ 2.01 ng/ml for controls; 4.4 _+ 2.6 for pts; morning: 14.9 _+ 5.3 for controls; 12.9 _+ 5.3 for pts), and did not report increased stress levels. There were no objective abnormalities in sleep architecture as measuredwith PSG in either patient group: sleep stage distribution, number of arousals were no different from controls. CONCLUSIONS:(1) IBS patients with dyspeptic symptoms report more severeand widespreadGI symptoms, but also more extraintestinslsymptoms including increasedsleep problems and greater psychopathology. (2) In the absence of objective sleep abnormalities, IBS patients do have sleep-related GI complaints. (3) Sleep complaints are not related to abnormal circadian changes in curt/sol Secretion or increasedstress levels in any 18S subpopulation. Instead, increasedsleep complaints may be attributableto a misperceptionof and/or a reporting bias regardingsleepquality.
613 Effects Of Stimulating Protease-Activated Rncoptors On Myatodc ~ in Guinea-Pig Small intestine Chuanyun Gag, Hong-Zhen Hu, Sumei L/u, Na Gag, Yun X/a, Jackie D. Wood, Ohio State Univ, Columbus, OH Background:Protease-activatedreceptors (PARs) are a family of G-protein coupled receptors that are cleaved and activated by certain proteases. Four members have been cloned and designated: PAR-l, 2, 3, 4. Among them, PAR-1 and PAR-2 have been localized in tissues of the gastrointestinal tract by immunohistochemistry and their roles in intestinal disorders assoclatad with inflammation have been implied. Nevertheless,the exact mechanism of an'don has not been fully clarified. This study tested the hypothesis that proteases modulate the gastrointestinalsystemthrough actions on the enteric nervous system. Methods: Conventional intracellular microelectrode and neuron tracer injection techniques were used to study the action of thrombin, trypsin, tryptase, PAR-1 tethered ligand analogue SFLLRL, and PAR-2 tethered ligand analogue SRLIGL on myenteric neurons of the guinea-pig small intestine. Results: Bath application of thrombin (O.l-lOunits/lOml) or trypsin (1-100 nM) evoked s tetrodoxin-resistant,concentration-dependentdepolarizingresponsesin meet of the myenteric neurons examined (77% in 56 S-type neurons, and 75% of 89 AH-type neurons). The depolarizationwas accompaniedby decreasedinput resistanceand associatedwith increased excitability reflected by repetitive spike discharge during depolarizing current pulses and anode-breakexcitation at the offset of hyperpolarizingcurrent pulses. Application of SFLLRI., the PAR-1 agonist, mimicked the effect of thrombin and trypsin with reduced potency of 0.11 p.M. In comparison,tryptase (0.1-10 nM) or PAR-2anon/st, SLIGRL, only produced a weak depolarizationof 2-4mV. The cyclooxygenaseinhibitor piroxicam (60 mM) did not affect the depolarizing responses produced by thrombin, trypsin or SFLLRL. Conclusion: The results suggest that the excitatory actions of proteasesare mediated primarily by PAR-t. PAR-2 may also contribute to the excitatory action of proteases,however,to a lesser extent. Arachidonic acid metabolism is not involved in the protease-evokedactivation of enteric neurons. (Supported by NIH R01 DK37238
614 is Psychiatric Illness, Personality or Both a Predictor of IBS in Young Adults? Results from a Birth Cohort Study in New Zealand. Stuart C. Howell, Univ of Sydney, Sydney Australia; Richie Pouifon, Univ of Otugo, Ounedin New Zealand; Nicholas J. Talley, Univ of Sydney, Sydney Australia Background:The association of IBS with psychiatric disorder remains uncertain. Relationships have been reported in some cross-sechonal studies, however, these have failed to establish whether psychiatric disorder precedes a diagnosis of IBS or is a consequence of if. Few studies haveexaminedwhether psychiatric disorder is more important than personalityfactors in the etiology of IBS. Method: The data were collected from a 1972 birth cohort (Dunedin, NZ), which has beenfollowed through the first 26 years of life: 96% of the living cohort was assessed at the most recent follow-up (at age 26). IBS was diagnosed according to the Manning criteria, using symptom data collected at age 26. DSM-IIIR diagnostic information was collected at ages 18 and 21 years; a DSM diagnosis at both time points was used to define chronic psychiatric disorder (any), chronic depressive illness and chronic anxiety disorder. Personality was assessed at age 18 years using the Multidimensional Personality Questionnaire (MPQ). Results: Datawas availablefor 869 subjects (49.6% female). Of these, 145 (18.7%) met the Manning criteria for IBS. This was significantly more common among females compared to males (19.5% vs 13.9%; p=O.03), but not among individuals with a history of psychiatric disorder. The lack of effect for psychiatric disorder was evident for any
A-II4
disorder (p=0.72), and for depressive illness (p=0.10) and anxiety disorder (p~0.53). Of the ten MPQ scales considered, only one emerged as a significant predictor of a diagnosis of IBS: mean scores on the Social Potency scale were significantly higher among subjects with IBS when compared to normal controls (Mean: 42.2 vs 36.6; p =0.02). The difference remained significant after adjustment for subject gender (p=O.01). Conclusions: In young adults, IBS is relatedto personality,but not to a history of psychiatricdisorder. When compared to healthy controls, IBS patients were significantly more likely to report a socially 'forceful' or dominant personality type.
615 Ps~intric Oiagnmms in GeatroenteralOy: Validation of a SeHoReporl Instrument (PRIME-MO Patient Health Qumdioneaira), Epidomiolo0y and Recognition, Philippe Persoons, Koen Luyckx, Benjamin Fischler, KatholiekeUniv Leuven, Dept Lialsonpcyohiatne, Leuven Belgium PURPOSE: In Gastroenterology(GE) no serf-report instrument to assess Anxiety and Mood Disorders (AMD) exists. We validated the AMD modules of the PRIME-MD Patient Health Questionnaire (PHO; Spitzer et al., JAMA 1999), the first categorial, self-report measure of psychialdc disorders, in GEpatientsand assessedthe epidemiologyand recognitionby medical residents of AMD in GE inpatients. METHODS: 639 consecutive GE in- and outpatients completed the PHQ. 184 patients were interviewed by a mental health professional with the Mini International Neoropsychlatric Interview 5.O.O to assess criterion validity. Construct validity was assessedwith the SF36 (Ouality of Life), Hospital Anxiety and Depressionscale (HAl)S), Beck Depression Inventory 7-items (BDI-PC), health care use and disability days. A questionnaireon the presenceof AMD in 175 inpatients was completed by medical residents, blind for the PHQ. Inpatients (n=475) were divided into 3 groups: Functional Intestinal Disorder (FID;n=87), Inflammatory Bowel Disease (IBO;n=lOl), Other Organic Disorder (OOD;n=287). RESULTS: Sensitivity and specificity were respectively 80.0% and 98.1% (K = 0.81) for Major DepressiveDisorder (MDD), 83.0% and 91.5% (K = 0.74) for Any Anxiety and DepressiveOisorder (AADO), 77.1% and 91.9% (K= 0.69) for Any DepressiveOisorder (ADD), 70.8% and 94.2% (K = 0.65) for Any Anxiety Disorder (AAD). Construct validity was satisfying and operating characteristics were similar or better than for HADS and BDI-PC. Prevalenceof AADD,ADD, MDD, Other DepressiveDisorder (ODD), AAD, Panic Disorder and Other Anxiety Disorder in inpatients was respectively36.0%, 31.4%, 17.9%, 13.5%, 14.9%, 6.8% and 9.6%. In lid there were significantly higher prevalences than in IBO and OOD (except for ODD) and in IBO there were significantly higher prevalencesfor AAOOand MOD than in OOD. Recognition rates by medical residents of AADO, ADO, MOD, ODD and AAD were respectively 27.0%, 22.7%, 10.8%, 13.8% and 20.7%. Recognition for AADD was significantly better in RO than in ISO and GOD. CONCLUSION:AMD are highly prevalent in GE inpatlents, especially in FID and IBD. AMO are poorly recognized by medical residents, especially in OOD and IBD. The PHG appears to be a diagnostic instrument for MDD and a good screening instrument for ADO and AAD in GE patients. The systematic use of the PHQ couM improve the recognition of AMD on GE wards.
616 Ookavioral aM Pl~ological Sleep Characteristics in IBS Symptom Subgroups S/god Elsonbruch, Dept of Medical Psychology, Univ Clin of Essen, Essen Germany; Jennifer J. Thompson, Michael J. Harnish, William C. Orr, Lynn Institute for Heaithcare Research, Ok/shores C/t/, OK BACKGROUND:Although subjective sleep complaints are well-documented in patients with IBS, the presence of objective sleep abnormalities remains controversial. It also remains unclear whether sleep dysfunction is relatedto patients gastrointestinal (GI) symptoms or to psychological factors. The goal of this study was to investigate sleep characteristics in IBS symtpom subgroups. METHODS:31 female/BS pts were stratified into 15 pts with only bowel symptoms (IBSonly), and 16 pts with both lower and upperdyspeptic-likesymptoms (IBS+ D). 21 healthy females served as controls. For 4 consecutive days subjective sleep quality, insomnia symptoms, alertness, state anxiety, stress levels, and daytime and night time GI symptoms were assessed. Saliva samples were collected for curt/sol analysis. On night 4, subjects underwent polysomnographic(PSG) moniforJngfor an objectiveassessmentof sleep quality including arousals and respiratory parameters. RESULTS:Comparedto both IBS-only and controls, IBS + D reported greater difficulties falling asleep, and showed a greater global Pittsburgh Sleep Quality Index (PSQI) score indicating greater overall sleep problems (all p<.05). A greater percentageof IBS + Opts reported night time GI symptoms (73.5 vs. 38.25 %, p<.01). Compared to both controls and IBS-only, IBS+D reported greater depression and morning anxiety across the 4 study (lays (all p<.05). Both patient groups felt less rested and showed decreasedlevels of daytime alertness on all 4 days compared to controls (all p<.01). Both patient groups showed normal circadian changes in salivary curt/sol levels (evening: 4.2 _+ 2.01 ng/ml for controls; 4.4 _+ 2.6 for pts; morning: 14.9 _+ 5.3 for controls; 12.9 _+ 5.3 for pts), and did not report increased stress levels. There were no objective abnormalities in sleep architecture as measuredwith PSG in either patient group: sleep stage distribution, number of arousals were no different from controls. CONCLUSIONS:(1) IBS patients with dyspeptic symptoms report more severeand widespreadGI symptoms, but also more extraintestinslsymptoms including increasedsleep problems and greater psychopathology. (2) In the absence of objective sleep abnormalities, IBS patients do have sleep-related GI complaints. (3) Sleep complaints are not related to abnormal circadian changes in curt/sol Secretion or increasedstress levels in any 18S subpopulation. Instead, increasedsleep complaints may be attributableto a misperceptionof and/or a reporting bias regardingsleepquality.