AGA Abstracts vs. 60.6 ± 6.5 pA (n=18, HFF serum + ZA)) and significantly increased number of APs at twice rheobase (1.5 ± 0.1 (n=22, HFF serum only) vs. 3.1 ± 0.4 (n=18, HFF serum + ZA, ***p=0.0009)). There were no differences when LFF serum was incubated with or without ZA. (P>0.05, all). ZA also prevented the inhibitory effect of Leptin-incubation. ZA+Leptin treatment significantly decreased rheobase (100.8 ± 18.6 pA (n=12, Leptin only) vs. 57.5 ± 5.8 pA (n=18, ZA+Leptin, *p=0.0193)) and increased action potential number (1.6 ± 0.3 (n=12, Leptin only) vs. 3.6 ± 0.7 (n=18, ZA+Leptin, *p=0.0277)). Leptin markedly attenuated Ca2+-signal in response to CCK and 5-HT. ZA significantly increased responses to CCK and 5-HT. (Control; 38.5 %( CCK, n=96), 36.4 %( 5-HT, n=66) vs. ZA; 67.6% (CCK, n= 74, ***p=0.0002), 68.1 %( 5-HT, n=88, ***p=0.0001)). STAT-3 inhibition with S3I-201 mimicked the effects of HFF serum, attenuating vagal afferent excitability. S3I-201 signifi- cantly increased rheobase (63.3 ± 6.1 pA (n=12, control) vs. 101.0 ±15.1 pA (n=10, S3I- 201)*p=0.0231). Conclusion: Inhibition of SOCS-3 prevented the inhibitory effects of leptin and obese mouse serum on the excitability and response to satiety mediators in vagal afferents. We suggest STAT-3/SOCS-3 pathway may play a key role in the defects in peripheral satiety signalling in obesity.(Funded by CIHR) Tu1784 Impaired Satiety Effect of GLP-1 Agonist in Obesity Is Associated With GLP-1 Resistance in Intestinal Afferent Nerves Alaa Al-Helaili, Beini Wang, Michael Beyak Intestinal afferent responses to satiety related stimuli are impaired in obesity. Glucagon Like Peptide-1 (GLP-1) is a satiety and incretin hormone that is released in response to luminal nutrients. The actions of peripheral GLP-1 may occur in part through vagal afferents. We hypothesize that GLP-1 receptor-mediated satiety and incretin effects are impaired in DIO mice, and that this is due to impaired sensitivity of intestinal afferents. Methods: Obesity was induced by feeding C57Bl6J mice a high fat diet for 10-12 weeks (HFF). LFF controls received a low fat (10% kcal from fat) diet (LFF). Food intake was monitored in LFF and HFF mice after i.p. injection of the stable GLP-1 agonist Exendin-4 (Ex-4)(100nmol/kg) or saline. Glucose tolerance tests were performed by gavaging mice with glucose, and measuring blood glucose concentrations every 15 min. In vitro extracellular intestinal afferent nerve recordings were performed using isolated segments of ileum from LFF and HFF mice. GLP- 1 (100nM) and Ex-4 (10nM) were perfused in the bath and intraluminally. Change in basal firing rate and response to distension were examined. Results: Ex-4 significantly decreased food intake in LFF mice compared to saline (4.5±0.6 vs. 2.7±0.2 Kcal / 3hours P≤0.03*). This decrease in food intake was not seen in HFF mice (2.6±0.4 vs. 2.2±0.5 Kcal /3hours P>0.05). HFF mice showed a significantly higher peak glucose compared to LFF mice (p<0.01). Ex-4 markedly attenuated the rise in blood glucose in both LFF and HFF mice, however the glucose at 30 min was slightly higher in the Ex-4 treated HFF mice compared to LFF (6.4±0.28 vs. 7.7±0.63 mmol/L p<0.05). LFF mice showed a significant increase in nerve firing (spike/s) in response to GLP-1 and Ex-4 (14.6±1.5 P≤0.0002) (10.4±2.6 P≤0.02) compared to baseline firing (3.6±1.1 vs 3±0.7) respectively. On the other hand, intestinal afferent firing rate was not significantly increased in HFF mice in response to GLP-1 and Ex-4 (13±5 P=0.2 vs 14.17±3.9 P=0.3) compared to baseline firing (6.1±1.3 vs 9.5±3.1) respectively. GLP-1 increased afferent response to distention at low pressures (15 mmHg) in LFF mice (10±5.5 vs. 23.3±4.4 P≤0.001, but not in HFF mice (12.2±4.9 vs. 15.8±6.5 P>0.05). Similarly, exendin-4 significantly increased afferent response to distention at low pressure (15 mmHg) in LFF mice (9.6±4.8 vs. 27±7.3 P≤0.001**) but not in DIO mice (18.6±4.4 vs. 19±5 P>0.05). Conclusion: Satiety effects of GLP-1 receptor activation are impaired in obesity. This is likely due to resistance to the effects of GLP-1 receptor activation at the level of the intestinal vagal afferent nerve terminal. The incretin effect of Ex-4 is preserved, with only a slightly higher post-prandial glucose in HFF mice. Loss of vagal afferent response to GLP-1 receptor stimulation may contribute to impaired control of food intake. (Funded by CIHR) Tu1785 Glucose Modulates Transmission From the Central Terminals of Vagal Afferents via a PKC-Dependent Pathway Ruchi Bhagat, Kirsteen N. Browning Ingested glucose causes gastric relaxation via activation of a vago-vagal reflex pathway. While these gastroinhibitory actions of glucose occur, in part, via a paracrine mechanism of action subsequent to the release of 5-HT from enteroendocrine cells, we have demonstrated previously that glucose also acts centrally to modulate the release of glutamate from the central terminals of vagal afferents via actions involving trafficking of 5-HT3 receptors. The aim of the present study was to determine the mechanism of action by which glucose regulates 5-HT3 receptor function on the central terminals of vagal afferents. Whole cell patch clamp recordings were made from nucleus of the tractus solitarius subnucleus centralis (NTScen) neurons in thin (300μm) rat brainstem slices. The ability of glucose (20mM) to modulate the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) was assessed in the presence and absence of protein kinase C (PKC) activators and inhibitors. The PKC activator, PMA (10nM) increased the frequency (180±38%; P<0.05) but not amplitude (101±5%; P>0.05) of mEPSCs in 13/14 neurons tested. In 8 neurons in which PMA increased mEPSC frequency (151±19% of control, P<0.05), increasing extracellu- lar glucose concentration from 10 to 20mM also increased mEPSC frequency (181±27%, P<0.05). In the presence of PMA, however, elevating extracellular glucose concentration had no further effect to increase mEPSC frequency (90±13%, P>0.05) suggesting PMA occluded the actions of glucose. The 5-HT3 receptor selective antagonist, ondansetron (1μM) prevented the ability of PMA to increase mEPSC frequency in 4/5 neurons tested (135±10% vs 93±14% in the presence of ondansetron, P>0.05). Additionally, in 6/8 neurons in which elevating extracellular glucose increased mEPSC frequency (173±16%, P<0.05), the PKCβ blocker CGP55353 (4μM) not only attenuated the facilitatory actions of glucose, but uncov- ered a presynaptic inhibitory effect of glucose (71±11%; P<0.05). These results suggest that glucose increases glutamatergic synaptic transmission from the central terminals of vagal afferents via a presynaptic PKCβ-dependent pathway to traffic 5-HT3 receptors. The uncover- ing of a presynaptic inhibitory action suggests the presence of a PKCβ-independent pathway S-842 AGA Abstracts by which glucose may additionally modulate vagal afferent transmission. The overall effect of glucose to regulate brainstem vagal neurocircuits, hence vagal efferent outflow to the viscera, may be dependent upon a balance of presynaptic facilitatory and inhibitory actions. Supported by NIH DK078364 Tu1786 Insulin Reverses the Diabetes-Induced Loss of Inhibitory Myenteric Neurons Luca Toti, Kirsteen N. Browning Upper gastrointestinal dysfunctions, including delayed gastric emptying, early satiety, nausea, bloating and abdominal pain, are experienced by a significant proportion of Type 1 diabetic patients. Several studies have indicated that some of this dysfunction may arise from a loss of inhibitory enteric neurons. While insulin replacement normalizes blood glucose levels, it is not known whether restored glycemic control is sufficient to attenuate or prevent the loss of inhibitory myenteric neurons. The aim of this study was to examine whether, in a rat model of Type 1 diabetes, glycemic reguluation (i) prevents the loss of inhibitory myenteric neurons (early insulin intervention) and (ii) reverses the loss of inhibitory enteric neurons (later insulin intervention). Male sprague-Dawley rats (~125-150g) were made diabetic via streptozotocin injection (STZ; 65mg/kg i.p.). Rats were considered to be hyperglycemic if blood glucose levels were above 250mg/dl. Diabetic rats received either no insulin (STZ group) or were implanted with a subcutaneous insulin pellet (26mg) after either 1 week (early intervention) or 6 weeks (late intervention) of uncontrolled hyperglycemia. After 12 weeks, rats were euthanized and their upper gastrointestinal tracts removed for fixation and subsequent immunohistochemical processing. Fundus, corpus and duodenum myenteric plexus preparations were stained for nitric oxide synthase immunoreactivity (NOS-IR); NOS- IR neurons were expressed as a percentage of total neuronal number of neurons, as revealed by PGP9.5 staining. In control rats, NOS-IR neurons constituted 35.4±1.0, 34.1±2.0 and 37.4±2.1% of neurons in fundus, corpus and duodenum, respectively (n=4 for each). As described previously, STZ-induced diabetes decreased the number of inhibitory NOS-IR myenteric neurons (24.0±2.6, 20.4±1.4 and 20.6±5.7% of neurons in fundus, corpus and duodenum, respectively; n=4; P<0.05 vs control). Insulin replacement after 1 week and 6 weeks restored the proportion of NOS-IR neurons to 31.8±2.2, 31.8±2.4 and 34.9±4.4% and 32.5±4.2, 32.9±3.2 and 33.0±3.7%, for fundus, corpus and duodenum, respectively (n=4 for each, P<0.05 vs STZ, P>0.05 vs control). These results demonstrate that (i) early insulin therapy prevents and (ii) late insulin therapy reverses the diabetes-induced loss of NOS-IR myenteric neurons. These results suggest that insulin may reverse the hyperglycemia- and diabetes-induced gastrointestinal dysfunction, in part, by restoring inhibitory neural regulation of the enteric nervous system. Supported by NIH DK078364 Tu1787 Role of Metabotropic Glutamate Receptors in Post-ERCP Model of Acute Pancreatitis Tanja Babic, R. Alberto Travagli Endoscopic retrograde cholangiopancreatography (ERCP) is a procedure used in the investi- gation and treatment of biliopancreatic disorders and is associated with high rates of acute pancreatitis. Pancreatic exocrine secretions (PES) are controlled by neurons in the dorsal motor nucleus of the vagus (DMV), whose activity is modulated by group II metabotropic glutamate receptors (mGluRs). We have recently shown that caerulein model of acute pancreatitis increases excitatory inputs to pancreas-projecting DMV neurons by decreasing the sensitivity of glutamatergic synaptic terminals to group II mGluR agonist APDC. The aim of this study was to determine if post-ERCP pancreatitis (PEP) alters the sensitivity of pancreas-projecting DMV neurons to APDC. PEP was induced in Sprague-Dawley rats by mechanical stimulation of the papilla of Vater. Presence of pancreatitis was confirmed by elevated plasma amylase levels. Experiments were conducted 18-24 hours after the induction of pancreatitis. In an in vivo anesthetized preparation, PES was collected via insertion of a cannula in the main pancreatic duct and collected as 10min samples for 30min before (baseline) and for 90min after the microinjection. The peak PES output was calculated as the average of the 3 samples after the microinjection. Microinjections of APDC (600pmoles/ 60nl) into the DMV of sham-operated animals elicited a significant increase in PES (140.1±8% baseline PES, p<0.05; n=3). Conversely, the same dose of APDC did not elicit a change in PEP (p>0.05; n=6). A separate group of animals received injections of 200μL of lidocaine into the pancreatic duct prior to ERCP. In these animals, APDC microinjections increased PES to the same degree as in sham-operated animals (140.0±7% baseline PES; p<0.05; n= 6). Whole-cell patch-clamp recordings were made from pancreas-projecting DMV neurons identified after placement of fluorescent dye on the pancreas. In control animals, perfusion of the slices with APDC (0.1-300μM) decreased the frequency of miniature excitatory postsynaptic currents (mEPSCs) in a dose-dependent manner. Perfusion of the slices with 100μM APDC decreased mEPSC frequency from 2.0±0.5 to 0.7±0.2 events/s (to 54.9±5% baseline; p<0.05; n=4). In PEP, perfusion of slices with 100μM APDC resulted in a smaller decrease in mEPSC frequency (from 5.1±3 events/s to 2.7±1.9 events/s; 46.2±9% of baseline; n=3). These data demonstrate that 1) PEP decreases the sensitivity of pancreas-projecting DMV neurons to APDC; 2) PEP induces similar changes in group II mGluRs on vagal circuits regulating PES as caerulein-induced model of acute pancreatitis; 3) intraductal lidocaine prevents PEP-induced vagal dysregulation. Supported by NIH DK55330 Tu1788 Sex Differences in Gabaergic Neurotransmission to Gastric-Projecting DMV Neurons Tanja Babic Women have a higher prevalence of functional gastrointestinal (GI) disorders and display reduced gastric motility compared to men. Gastric motility is modulated by neurons of the dorsal motor nucleus of the vagus (DMV) and the activity of these neurons is regulated by a robust tonic GABAergic input. In general, females have been shown to have a higher