24 Interdigestive Gastric Motility and Gastric Emptying Are No More Impaired Following Repeated...
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levels were determined by enzyme immunoassay. RESULTS: Intracerebroventricular adminis- tration of m-CPP and fenfluramine to fasted rats altered the fasted motor activities in the antrum and duodenum to fed-like motor activities and the phase III-like contractions were significantly decreased. Treatment with the 5-HT2cR antagonist (SB242084), but not the 5-HT1bR (SB224289) and 5-HT2bR (SB215505) antagonists, was able to restore the fasted motor patterns in the antrum and duodenum of fenfluramine-treated rats. SB242084 also inhibited the decrease in plasma acyl ghrelin induced by fenfluramine. The MC4R (HS014) and MC3/4R (SHU9119) antagonists failed to modify the decreased motor activities induced by fenfluramine. In contrast, ghrelin and NPY restored the fasted motor patterns in the antrum and duodenum of fenfluramine-treated rats. Administration of phentolamine and hesperidine, which stimulate the secretion of endogenous acyl ghrelin, successfully restored the fasted motor activities in fenfluramine-treated rats. CONCLUSION: The present study has elucidated a novel central 5-HT2c receptor pathway regulating physiological GI motor activity. These results demonstrate direct neuroendocrine control over GI function by central 5-HT2c receptors, which inhibit ghrelin-NPY signaling. 24 Interdigestive Gastric Motility and Gastric Emptying Are No More Impaired Following Repeated Restraint Stress in Rats; the Possible Involvement of Central CRF, Gastric Ghrelin and HPA Axis in the Adaptation Process to Chronic Stress Jun Zheng, Reji Babygirija, Anthony Dobner, Ludwig Kirk, Toku Takahashi Although chronic stress is highly associated with GI motor disorders, effects of chronic stress on gastric motility have not been fully studied. We compared the effects of acute and chronic stress on solid gastric emptying and interdigestive gastroduodenal contractions in rats. SD rats were loaded with single restraint stress (acute stress) or repeated restraint stress for 5 consecutive days (chronic stress). To investigate whether gastric motility responses to central corticotropin releasing factor (CRF) are altered following chronic stress, CRF (1 μg) was administered intracisternally (ic). To evaluate the hypothalamus-pituitary-adrenal (HPA) axis, plasma corticosterone levels were measured. The changes of plasma levels and gastric mRNA expression of ghrelin were also studied following chronic stress. In non-restraint rats, solid gastric emptying was 52.4 +/- 3.1% 90 min after the feeding of 1.6 g rat chow. Acute stress significantly delayed gastric emptying (25.6 +/- 3.9%), which was completely restored (53.3 +/- 3.6%) at the day 5 of consecutive stress loading. Appearance of gastric, but not duodenal, phase III-like contractions was completely abolished during acute stress. Reduced frequency of gastric phase III-like contractions in response to restraint stress was gradually increased after day 3 and appearance of phase III-like contractions was completely restored on day 5. IC-injection of CRF significantly delayed solid gastric emptying and completely abolished gastric phase III-like contractions in non-restraint rats as well as chronically restraint rats. No significant differences of the plasma levels of active ghrelin were observed between control rats and restraint rats at day 1 (21.8 +/- 1.9 pg/ml). In contrast, plasma ghrelin levels were significantly increased to 65.6 +/- 5.4 pg/ml at day 5. Ghrelin mRNA was also significantly increased to 143 +/- 12% of controls at day 5. Plasma corticosterone levels significantly increased to 1315.3 +/- 220.4 ng/ml in response to acute stress, compared to that of non-restraint rats (70.2 +/- 12.0 ng/ml). At the day 5, there was no more significant increase observed in plasma corticosterone levels. Our current study shows that acute restraint stress-induced impaired gastric motility was completely restored following chronic stress. It is suggested that attenuated HPA axis and upregulated ghrelin expression are involved in the adaptation process following chronic stress. The inhibitory pathway of gastric motility in response to central CRF is not altered following chronic stress. It is conceivable that chronic stress attenuates CRF expression at the CNS, resulting in restored gastric motility. 25 Supraspinal Activation Associated with Stress-Induced Visceral Hypersensitivity: A Neuroimaging Study in the Awake Rat Pamela E. Paulson, Thomas J. Morrow, John W. Wiley, Shuangsong Hong Background: Stress-induced visceral hypersensitivity has been suggested as an important pathophysiological mechanism of irritable bowel syndrome (IBS). However, the underlying mechanism(s) are not fully understood. Pain perception is affected by both peripheral and central alterations. Recently, we reported that visceral hyperalgesia in response to colorectal distention (CRD) in the repeated water avoidance (WA)-stressed rat was associated with altered endocannabinoid and TRPV1 receptor expression in dorsal root ganglia neurons. In the present study, we examined visceral sensitivity response to colorectal distension (CRD) combined with brain imaging to identify potential central mechanisms involved in stress- induced visceral hyperalgesia. Methods: Male rats were exposed to 1-hour WA stress daily for 10 consecutive days. The visceromotor response (VMR) to CRD was electromyographically measured at distension pressures of 10, 20, 40, and 60 mmHg. On day 11, neuroimaging studies were performed in freely moving, awake rats during a 60 second colorectal distension (balloon distension) at 40 mmHg pressure. Neuronal activation in thirty-seven brain regions was determined by autoradiographic measurement of regional cerebral blood flow (rCBF) using the radiotracer [99m]Tc-HMPAO. Repeated measures ANOVA with Dunnett's T3 for post-hoc analysis was used to test for significant differences in the activation index (AI) for each region of interest (ROI) between stressed and control rats. Results: WA stressed rats demonstrated a significant increase in VMR at distension pressures of 40 and 60 mmHg compared to controls. The WA stressed rats showed significant increases in brain activation in regions involved in pain processing following colorectal distension (40 mmHg) compared to stimulated controls. Specifically, we found increased activation in several thalamic regions, including the ventral anterior and ventral posteromedial nuclei, the medial thalamus and posterior thalamic groups. In addition, significant AI increases following CRD were found in multiple limbic regions implicated in antinociception including the nucleus accumbens, substantia nigra, subthalamic nucleus, the arcuate nucleus, habenular complex, periaqueduc- tal gray and the prelimbic cortex. Finally, activation in the the paraventricular nucleus (PVN) of the hypothalamus was enhanced in the WA stress group. Conclusion: Visceral hyperalgesia induced by WA stress is associated with enhanced brain activation in response to colorectal A-3 AGA Abstracts distension. These data provide a basis for further studies to improve our understanding of brain-gut interactions and the enhanced visceral hypersensitivity in IBS patients. 26 Accurate CA 2+ Buffering and Neurotransmitter Release Requires Functional Mitochondria At Enteric Varicosities and Release Sites Pieter Vanden Berghe, Werend Boesmans, Valentine Van den Abbeel, Jan F. Tack Mitochondria are vital organelles that produce energy through oxidative phosphorylation. They are positioned at sites of high ATP consumption for instance at neurotransmitter release sites, where they, apart from generating energy, also can take part in buffering intracellular Ca 2+ . A side effect of producing ATP is possible leakage of harmful agents such as superoxides, rendering these organelles especially sensitive to oxidative pressure. The aim of this study was to investigate the role of mitochondria and the antioxidant ascorbic acid (vitamin C) in local Ca 2+ buffering and neurotransmitter release at enteric varicosities. Cultured guinea- pig myenteric neurons were used, which we loaded with Fluo-4 to measure cytosolic Ca 2+ signals in terminals and varicosities or stained with FM1-43 to directly monitor synaptic vesicle release. We also used transgenic synaptopHluorin (sypH) mice expressing a pH- sensitive GFP-reporter that switches on during neurotransmission. Electrical stimuli were applied with a miniature bipolar electrode positioned over the microscopic field of view and fluorescent signals were analyzed using Igor Pro®. Acute application of carbonylcyanide- p-trifluoromethoxyphenylhydrazone (FCCP, 10 -6 M) was used to uncouple oxidative phos- phorylation and impair mitochondrial function. Ascorbic acid (10 -5 M) was either adminis- tered acutely or 1 day prior to the recordings. Brief electrical stimuli (0.5s, 20Hz) elicited highly repeatable transient Ca 2+ responses in nerve varicosities and boutons. Acute impair- ment of mitochondrial function slightly reduced response amplitude (1.19±0.01 vs. 1.26±0.01, p=0.01, n=41) but more importantly it prolonged its duration by approximately 50% (4.6±0.4 vs. 3.4±0.08 s, p<0.001, n=41). Preincubation with ascorbic acid, partially countered the broadening effect of FCCP (4.2±0.1 vs 4.6±0.4 s, p<0.02, n=30), although there was no acute effect of ascorbic acid administration. Active release sites, stained with FM1-43, released a significantly larger fraction of vesicles after a 2s stimulus (20Hz) in the presence of FCCP compared to control (38±2 vs. 19±1%, p<0.01). Also in enteric release sites of sypH mice, FCCP caused a marked (15%) increase in fluorescence, indicative of massive neurotransmitter release. Conclusion: Mitochondrial Ca 2+ buffering is essential to accurately control transmitter release. Defective mitochondria lead to exaggerated release due to impaired Ca 2+ buffering, which can be partially reversed by ascorbic acid pretreatment. Impairment of mitochondria may be the mechanism underlying enhanced neurotransmitter release as observed in experimental models of inflammation. 27 Novel Purinergic P1, P2X and P2Y Receptor Signaling Targets in the Human Enteric Nervous System and Glia Jacqueline E. Wunderlich, Tianhua Ren, Bradley Needleman, Dean J. Mikami, Iveta Grants, Fievos L. Christofi Background/Aim: Little or no information exists on synaptic transmission and modulation or purinergic transmission in human enteric nervous system (huENS;AJP,294:G554- G566,2008). Most of our knowledge is from rodent species. Our goal was to assess the role of purinergic transmission and signaling in huENS and ganglia. Methods: Microdissected intestinal submucosa from Roux-en-Y patients were loaded with 30μM Fluo-4/AM Ca 2+ and time-series analysis of intracellular Ca 2+ was done using laser confocal imaging. TTX-sensitive synaptic Ca 2+ responses were elicited by focal electrical stimulation of fiber tracts (FTS) 0.3mm away from a recording ganglion (G); imaging was done at slow (1-2s/frame) or fast rate (100msec/ROI of 8x8μm 2 ). Results: Ca 2+ analysis was done in 16 Roux-en-Y surgical cases, 65 ganglia, 420 neurons and 100 glia. A 3s FTS elicited frequency-dependent synaptic Ca 2+ responses at 0.1Hz-100Hz with single or multiple fast and slow components. Apyrase (1.5-8U/ml) suppressed synaptic responses by 69%±7% (p=4.0e -6 ;3G,n=23/28). The ecto- ATPase inhibitor ARL 67156 augmented FTS responses by 14%±4% (p=0.0177,n=7). Aden- osine deaminase (0.5-2 U/ml) augmented peak FTS responses by 59±8% (p=2.5e -10 ;5G,n= 27/72) and adenosine inhibited them (1nM-1mM,p<0.0001,n=152) with IC 50 =25μM and 58±7% maximum inhibition (p=6.9e -5 ;5G,n=29,p=6.9e -5 ). A P2X(2/3/4) receptor (R) antag- onist 2'3'-O-trinitrophenyl-ATP (TNP-ATP,20nM) reduced FTS responses by 49±9% (p= 0.005;10G,n=15/64);5μM TNP-ATP reduced responses in different neurons by 54±4% (n= 31/56; p=5e -10 ,10G). Strong P2X3 immunoreactivity (ir) occurred in < 3% of neurons. The P2Y agonist adenosine-5'-O-(2-thiodiphosphate)(ADPβS,5μM) reduced FTS Ca 2+ responses by 66±5% (p<0.0001,n=41,5G). Some FTS Ca 2+ responses were sensitive to blockade with 5μM TNP-ATP and ADPβS, respectively (n=8/25,3G). Sequential perfusion of a P2X (αβMe- ATP,5μM), a P2Y (ADPβS,) or P2Y14 agonist (UDP-glucose,2μM) differentially evoked complex and often cyclic Ca 2+ transients in neurons or glia; subsets of cells have one or more receptor responses (i.e.P2Y/P2Y14/P2X;P2Y only). P2Y12R-ir was abundant in submucous neurons while P2Y2R-ir was rare in neurons but more expressed in glia. FTS, a nicotinic agonist DMPP (100μM) or high potassium depolarization elicited Ca 2+ responses in neurons and a delayed response in glia - Apyrase or TTX could abolish glial responses. Conclusion: Novel data suggest that purines play an important role in synaptic transmission and neural-glial communication in huENS. Putative functional targets include P1, P2X and P2Y receptors that are differentially expressed in subsets of neurons or glia (DK44179, NCRR 1S10RR11434). 28 Cannabinoids Are Retrograde Transmitters in the Myenteric Plexus Ian M. Hons, Beat Lutz, Keith A. Sharkey Background: The endocannabinoid (eCB) signalling system, including cannabinoid (CB) receptors and the enzymes for the synthesis and degradation of eCBs, has been identified in the enteric nervous sysem (ENS) of the mouse. In mice deficient in the CB1 receptor AGA Abstracts
Transcript of 24 Interdigestive Gastric Motility and Gastric Emptying Are No More Impaired Following Repeated...
levels were determined by enzyme immunoassay. RESULTS: Intracerebroventricular adminis-tration of m-CPP and fenfluramine to fasted rats altered the fasted motor activities in theantrum and duodenum to fed-like motor activities and the phase III-like contractions weresignificantly decreased. Treatment with the 5-HT2cR antagonist (SB242084), but not the5-HT1bR (SB224289) and 5-HT2bR (SB215505) antagonists, was able to restore the fastedmotor patterns in the antrum and duodenum of fenfluramine-treated rats. SB242084 alsoinhibited the decrease in plasma acyl ghrelin induced by fenfluramine. The MC4R (HS014)and MC3/4R (SHU9119) antagonists failed to modify the decreased motor activities inducedby fenfluramine. In contrast, ghrelin and NPY restored the fasted motor patterns in theantrum and duodenum of fenfluramine-treated rats. Administration of phentolamine andhesperidine, which stimulate the secretion of endogenous acyl ghrelin, successfully restoredthe fasted motor activities in fenfluramine-treated rats. CONCLUSION: The present studyhas elucidated a novel central 5-HT2c receptor pathway regulating physiological GI motoractivity. These results demonstrate direct neuroendocrine control over GI function by central5-HT2c receptors, which inhibit ghrelin-NPY signaling.
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Interdigestive Gastric Motility and Gastric Emptying Are No More ImpairedFollowing Repeated Restraint Stress in Rats; the Possible Involvement ofCentral CRF, Gastric Ghrelin and HPA Axis in the Adaptation Process toChronic StressJun Zheng, Reji Babygirija, Anthony Dobner, Ludwig Kirk, Toku Takahashi
Although chronic stress is highly associated with GI motor disorders, effects of chronicstress on gastric motility have not been fully studied. We compared the effects of acute andchronic stress on solid gastric emptying and interdigestive gastroduodenal contractions inrats. SD rats were loaded with single restraint stress (acute stress) or repeated restraint stressfor 5 consecutive days (chronic stress). To investigate whether gastric motility responses tocentral corticotropin releasing factor (CRF) are altered following chronic stress, CRF (1 μg)was administered intracisternally (ic). To evaluate the hypothalamus-pituitary-adrenal (HPA)axis, plasma corticosterone levels were measured. The changes of plasma levels and gastricmRNA expression of ghrelin were also studied following chronic stress. In non-restraintrats, solid gastric emptying was 52.4 +/- 3.1% 90 min after the feeding of 1.6 g rat chow.Acute stress significantly delayed gastric emptying (25.6 +/- 3.9%), which was completelyrestored (53.3 +/- 3.6%) at the day 5 of consecutive stress loading. Appearance of gastric,but not duodenal, phase III-like contractions was completely abolished during acute stress.Reduced frequency of gastric phase III-like contractions in response to restraint stress wasgradually increased after day 3 and appearance of phase III-like contractions was completelyrestored on day 5. IC-injection of CRF significantly delayed solid gastric emptying andcompletely abolished gastric phase III-like contractions in non-restraint rats as well aschronically restraint rats. No significant differences of the plasma levels of active ghrelinwere observed between control rats and restraint rats at day 1 (21.8 +/- 1.9 pg/ml). Incontrast, plasma ghrelin levels were significantly increased to 65.6 +/- 5.4 pg/ml at day 5.Ghrelin mRNA was also significantly increased to 143 +/- 12% of controls at day 5. Plasmacorticosterone levels significantly increased to 1315.3 +/- 220.4 ng/ml in response to acutestress, compared to that of non-restraint rats (70.2 +/- 12.0 ng/ml). At the day 5, there wasno more significant increase observed in plasma corticosterone levels. Our current studyshows that acute restraint stress-induced impaired gastric motility was completely restoredfollowing chronic stress. It is suggested that attenuated HPA axis and upregulated ghrelinexpression are involved in the adaptation process following chronic stress. The inhibitorypathway of gastric motility in response to central CRF is not altered following chronic stress.It is conceivable that chronic stress attenuates CRF expression at the CNS, resulting inrestored gastric motility.
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Supraspinal Activation Associated with Stress-Induced VisceralHypersensitivity: A Neuroimaging Study in the Awake RatPamela E. Paulson, Thomas J. Morrow, John W. Wiley, Shuangsong Hong
Background: Stress-induced visceral hypersensitivity has been suggested as an importantpathophysiological mechanism of irritable bowel syndrome (IBS). However, the underlyingmechanism(s) are not fully understood. Pain perception is affected by both peripheral andcentral alterations. Recently, we reported that visceral hyperalgesia in response to colorectaldistention (CRD) in the repeated water avoidance (WA)-stressed rat was associated withaltered endocannabinoid and TRPV1 receptor expression in dorsal root ganglia neurons. Inthe present study, we examined visceral sensitivity response to colorectal distension (CRD)combined with brain imaging to identify potential central mechanisms involved in stress-induced visceral hyperalgesia. Methods: Male rats were exposed to 1-hour WA stress dailyfor 10 consecutive days. The visceromotor response (VMR) to CRDwas electromyographicallymeasured at distension pressures of 10, 20, 40, and 60 mmHg. On day 11, neuroimagingstudies were performed in freely moving, awake rats during a 60 second colorectal distension(balloon distension) at 40 mmHg pressure. Neuronal activation in thirty-seven brain regionswas determined by autoradiographic measurement of regional cerebral blood flow (rCBF)using the radiotracer [99m]Tc-HMPAO. Repeated measures ANOVA with Dunnett's T3 forpost-hoc analysis was used to test for significant differences in the activation index (AI) foreach region of interest (ROI) between stressed and control rats. Results: WA stressed ratsdemonstrated a significant increase in VMR at distension pressures of 40 and 60 mmHgcompared to controls. The WA stressed rats showed significant increases in brain activationin regions involved in pain processing following colorectal distension (40 mmHg) comparedto stimulated controls. Specifically, we found increased activation in several thalamic regions,including the ventral anterior and ventral posteromedial nuclei, the medial thalamus andposterior thalamic groups. In addition, significant AI increases following CRD were foundin multiple limbic regions implicated in antinociception including the nucleus accumbens,substantia nigra, subthalamic nucleus, the arcuate nucleus, habenular complex, periaqueduc-tal gray and the prelimbic cortex. Finally, activation in the the paraventricular nucleus (PVN)of the hypothalamus was enhanced in theWA stress group.Conclusion: Visceral hyperalgesiainduced by WA stress is associated with enhanced brain activation in response to colorectal
A-3 AGA Abstracts
distension. These data provide a basis for further studies to improve our understanding ofbrain-gut interactions and the enhanced visceral hypersensitivity in IBS patients.
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Accurate CA2+ Buffering and Neurotransmitter Release Requires FunctionalMitochondria At Enteric Varicosities and Release SitesPieter Vanden Berghe, Werend Boesmans, Valentine Van den Abbeel, Jan F. Tack
Mitochondria are vital organelles that produce energy through oxidative phosphorylation.They are positioned at sites of high ATP consumption for instance at neurotransmitter releasesites, where they, apart from generating energy, also can take part in buffering intracellularCa2+. A side effect of producing ATP is possible leakage of harmful agents such as superoxides,rendering these organelles especially sensitive to oxidative pressure. The aim of this studywas to investigate the role of mitochondria and the antioxidant ascorbic acid (vitamin C)in local Ca2+ buffering and neurotransmitter release at enteric varicosities. Cultured guinea-pig myenteric neurons were used, which we loaded with Fluo-4 to measure cytosolic Ca2+
signals in terminals and varicosities or stained with FM1-43 to directly monitor synapticvesicle release. We also used transgenic synaptopHluorin (sypH) mice expressing a pH-sensitive GFP-reporter that switches on during neurotransmission. Electrical stimuli wereapplied with a miniature bipolar electrode positioned over the microscopic field of viewand fluorescent signals were analyzed using Igor Pro®. Acute application of carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP, 10-6M) was used to uncouple oxidative phos-phorylation and impair mitochondrial function. Ascorbic acid (10-5M) was either adminis-tered acutely or 1 day prior to the recordings. Brief electrical stimuli (0.5s, 20Hz) elicitedhighly repeatable transient Ca2+ responses in nerve varicosities and boutons. Acute impair-ment of mitochondrial function slightly reduced response amplitude (1.19±0.01 vs.1.26±0.01, p=0.01, n=41) but more importantly it prolonged its duration by approximately50% (4.6±0.4 vs. 3.4±0.08 s, p<0.001, n=41). Preincubation with ascorbic acid, partiallycountered the broadening effect of FCCP (4.2±0.1 vs 4.6±0.4 s, p<0.02, n=30), althoughthere was no acute effect of ascorbic acid administration. Active release sites, stained withFM1-43, released a significantly larger fraction of vesicles after a 2s stimulus (20Hz) in thepresence of FCCP compared to control (38±2 vs. 19±1%, p<0.01). Also in enteric releasesites of sypH mice, FCCP caused a marked (15%) increase in fluorescence, indicative ofmassive neurotransmitter release. Conclusion: Mitochondrial Ca2+ buffering is essential toaccurately control transmitter release. Defective mitochondria lead to exaggerated releasedue to impaired Ca2+ buffering, which can be partially reversed by ascorbic acid pretreatment.Impairment of mitochondria may be the mechanism underlying enhanced neurotransmitterrelease as observed in experimental models of inflammation.
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Novel Purinergic P1, P2X and P2Y Receptor Signaling Targets in the HumanEnteric Nervous System and GliaJacqueline E. Wunderlich, Tianhua Ren, Bradley Needleman, Dean J. Mikami, IvetaGrants, Fievos L. Christofi
Background/Aim: Little or no information exists on synaptic transmission and modulationor purinergic transmission in human enteric nervous system (huENS;AJP,294:G554-G566,2008). Most of our knowledge is from rodent species. Our goal was to assess the roleof purinergic transmission and signaling in huENS and ganglia. Methods: Microdissectedintestinal submucosa from Roux-en-Y patients were loaded with 30μM Fluo-4/AM Ca2+ andtime-series analysis of intracellular Ca2+ was done using laser confocal imaging. TTX-sensitivesynaptic Ca2+ responses were elicited by focal electrical stimulation of fiber tracts (FTS)0.3mm away from a recording ganglion (G); imaging was done at slow (1-2s/frame) or fastrate (100msec/ROI of 8x8μm2). Results: Ca2+ analysis was done in 16 Roux-en-Y surgicalcases, 65 ganglia, 420 neurons and 100 glia. A 3s FTS elicited frequency-dependent synapticCa2+ responses at 0.1Hz-100Hz with single or multiple fast and slow components. Apyrase(1.5-8U/ml) suppressed synaptic responses by 69%±7% (p=4.0e-6;3G,n=23/28). The ecto-ATPase inhibitor ARL 67156 augmented FTS responses by 14%±4% (p=0.0177,n=7). Aden-osine deaminase (0.5-2 U/ml) augmented peak FTS responses by 59±8% (p=2.5e-10;5G,n=27/72) and adenosine inhibited them (1nM-1mM,p<0.0001,n=152) with IC50=25μM and58±7% maximum inhibition (p=6.9e-5;5G,n=29,p=6.9e-5). A P2X(2/3/4) receptor (R) antag-onist 2'3'-O-trinitrophenyl-ATP (TNP-ATP,20nM) reduced FTS responses by 49±9% (p=0.005;10G,n=15/64);5μM TNP-ATP reduced responses in different neurons by 54±4% (n=31/56; p=5e-10,10G). Strong P2X3 immunoreactivity (ir) occurred in < 3% of neurons. TheP2Y agonist adenosine-5'-O-(2-thiodiphosphate)(ADPβS,5μM) reduced FTS Ca2+ responsesby 66±5% (p<0.0001,n=41,5G). Some FTS Ca2+ responses were sensitive to blockade with5μM TNP-ATP and ADPβS, respectively (n=8/25,3G). Sequential perfusion of a P2X (αβMe-ATP,5μM), a P2Y (ADPβS,) or P2Y14 agonist (UDP-glucose,2μM) differentially evokedcomplex and often cyclic Ca2+ transients in neurons or glia; subsets of cells have oneor more receptor responses (i.e.P2Y/P2Y14/P2X;P2Y only). P2Y12R-ir was abundant insubmucous neurons while P2Y2R-ir was rare in neurons but more expressed in glia. FTS,a nicotinic agonist DMPP (100μM) or high potassium depolarization elicited Ca2+ responsesin neurons and a delayed response in glia - Apyrase or TTX could abolish glial responses.Conclusion: Novel data suggest that purines play an important role in synaptic transmissionand neural-glial communication in huENS. Putative functional targets include P1, P2X andP2Y receptors that are differentially expressed in subsets of neurons or glia (DK44179,NCRR 1S10RR11434).
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Cannabinoids Are Retrograde Transmitters in the Myenteric PlexusIan M. Hons, Beat Lutz, Keith A. Sharkey
Background: The endocannabinoid (eCB) signalling system, including cannabinoid (CB)receptors and the enzymes for the synthesis and degradation of eCBs, has been identifiedin the enteric nervous sysem (ENS) of the mouse. In mice deficient in the CB1 receptor
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