Expression of 5-HT3 receptors by extrinsic duodenal afferents contribute to intestinal inhibition of gastric emptying

Intestinal perfusion with carbohydrates inhibits gastric emptying via vagal and spinal capsaicin-sensitive afferent pathways. The aim of the present study was to determine the role of 1) 5-hydroxytryptamine (5-HT)(3) receptors (5-HT(3)R) in mediating glucose-induced inhibition of gastric emptying and 2) 5-HT(3)R expression in vagal and spinal afferents in innervating the duodenum. In awake rats fitted with gastric and duodenal cannulas, perfusion of the duodenum with glucose (50 and 100 mg) inhibited gastric emptying. Intestinal perfusion of mannitol inhibited gastric emptying only at the highest concentration (990 mosm/kgH(2)O). Pretreatment with the 5-HT(3)R antagonist tropisetron abolished both glucose- and mannitol-induced inhibition of gastric emptying. Retrograde labeling of visceral afferents by injection of dextran-conjugated Texas Red into the duodenal wall was used to identify extrinsic primary afferents. Immunoreactivity for 5-HT(3)R, visualized with an antibody directed to the COOH terminus of the rat 5-HT(3)R, was found in >80% of duodenal vagal and spinal afferents. These results show that duodenal extrinsic afferents express 5-HT(3)R and that the receptor mediates specific glucose-induced inhibition of gastric emptying. These findings support the hypothesis that enterochromaffin cells in the intestinal mucosa release 5-HT in response to glucose, which activates 5-HT(3)R on afferent nerve terminals to evoke reflex changes in gastric motility. The primary glucose sensors of the intestine may be mucosal enterochromaffin cells.     

Inhibition of gastric motility by hyperglycemia is mediated by nodose ganglia KATP channels

The inhibitory action of hyperglycemia is mediated by vagal afferent fibers innervating the stomach and duodenum. Our in vitro studies showed that a subset of nodose ganglia neurons is excited by rising ambient glucose, involving inactivation of ATP-sensitive K(+) (K(ATP)) channels and leading to membrane depolarization and neuronal firing. To investigate whether nodose ganglia K(ATP) channels mediate gastric relaxation induced by hyperglycemia, we performed in vivo gastric motility studies to examine the effects of K(ATP) channel activators and inactivators. Intravenous infusion of 20% dextrose induced gastric relaxation in a dose-dependent manner. This inhibitory effect of hyperglycemia was blocked by diazoxide, a K(ATP) channel activator. Conversely, tolbutamide, a K(ATP) channel inactivator, induced dose-dependent gastric relaxation, an effect similar to hyperglycemia. Vagotomy, perivagal capsaicin treatment, and hexamethonium each prevented the inhibitory action of tolbutamide. Similarly, N(G)-nitro-l-arginine methyl ester, an inhibitor of nitric oxide synthase, also blocked tolbutamide's inhibitory effect. To show that K(ATP) channel inactivation at the level of the nodose ganglia induces gastric relaxation, we performed electroporation of the nodose ganglia with small interfering RNA of Kir6.2 (a subunit of K(ATP)) and plasmid pEGFP-N1 carrying the green fluorescent protein gene. The gastric responses to hyperglycemia and tolbutamide were not observed in rats with Kir6.2 small interfering RNA-treated nodose ganglia. However, these rats responded to secretin, which acts via the vagal afferent pathway, independently of K(ATP) channels. These studies provide in vivo evidence that hyperglycemia induces gastric relaxation via the vagal afferent pathway. This action is mediated through inactivation of nodose ganglia K(ATP) channels.     

Glucose increases synaptic transmission from vagal afferent central nerve terminals via modulation of 5-HT3 receptors

Acute hyperglycemia has profound effects on vagally mediated gastrointestinal functions. We have reported recently that the release of glutamate from the central terminals of vagal afferent neurons is correlated directly with the extracellular glucose concentration. The present study was designed to test the hypothesis that 5-HT(3) receptors present on vagal afferent nerve terminals are involved in this glucose-dependent modulation of glutamatergic synaptic transmission. Whole-cell patch-clamp recordings were made from neurons of the nucleus tractus solitarius (NTS) in thin rat brainstem slices. Spontaneous and evoked glutamate release was decreased in a concentration-dependent manner by the 5-HT(3) receptor selective antagonist, ondansetron. Alterations in the extracellular glucose concentration induced parallel shifts in the ondansetron-mediated inhibition of glutamate release. The changes in excitatory synaptic transmission induced by extracellular glucose concentration were mimicked by the serotonin uptake inhibitor, fenfluramine. These data suggest that glucose alters excitatory synaptic transmission within the rat brainstem via actions on tonically active 5-HT(3) receptors, and the number of 5-HT(3) receptors on vagal afferent nerve terminals is positively correlated with the extracellular glucose concentration. These data indicate that the 5-HT(3) receptors present on synaptic connections between vagal afferent nerve terminals and NTS neurons are a strong candidate for consideration as one of the sites where glucose acts to modulate vagovagal reflexes.     

Capsaicin-sensitive vagal afferents and CCK in inhibition of gastric motor function induced by intestinal nutrients

The role of vagal afferent pathways and cholecystokinin (CCK) in mediating changes in gastric motor function after a meal was investigated in urethane-anesthetized rats. Proximal gastric motor function was measured manometrically, and nutrients were infused into an isolated segment of duodenum. Inhibition of gastric motility in response to duodenal infusion of protein (peptone or casein), but not carbohydrate (glucose), was significantly attenuated by administration of the CCK antagonist, L364,718. Selective ablation of vagal afferents by perineural treatment with the sensory neurotoxin, capsaicin, significantly reduced responses to both duodenal protein and glucose. These results suggest that protein in the duodenum decreases proximal gastric motor function via release of CCK and a vagal capsaicin-sensitive afferent pathway. In contrast, glucose acts via a capsaicin-sensitive vagal pathway not involving CCK. Thus separate neural and hormonal mechanisms mediate the effects of different nutrients in the duodenal feedback regulation of gastric motor function.     

Involvement of 5-HT(2) receptor in imipramine-induced hyperglycemia in mice.

Effects of imipramine on plasma glucose levels were investigated in mice. Imipramine i. p. induced dose-dependent hyperglycemia, which was enhanced by pretreatment with 5-HT (1/2/5/7) receptor antagonist methysergide and 5-HT (2A/2B/2C) receptor antagonist LY 53857. 5-HT (2C/2B) receptor antagonist SB 206553 also augmented imipramine-induced hyperglycemia although 5-HT (1A) and 5-HT (1B) receptor antagonist (-)-propranolol,5-HT (2A) receptor antagonist ketanserin and 5-HT (3/4) receptor antagonist tropisetron each had no effect. Imipramine i. p.-induced hyperglycemia was antagonized by the 5-HT (2C/2B) receptor agonist 1-(3-chlorophenyl)piperazine (mCPP), while the 5-HT (2B) receptor agonist BW 723C86 had no effect. Intracerebroventricular injection of imipramine also elevated plasma glucose levels, which is enhanced by SB 206553. Hyperglycemia elicited by central injection of imipramine was abolished by adrenalectomy. These results suggest that imipramine-induced hyperglycemia in mice is related to its inhibition of the central 5-HT (2C) receptor. Moreover, our results indicate that adrenaline release is related to imipramine-induced hyperglycemia.     

Receptor subtype specific activation of the rat gastric vagal afferent fibers to serotonin

Systemic administration (i.v.) of serotonin (5-HT) evoked a transient vagal afferent nerve discharge, bradycardia, and hypotension in the rat. The half-effective dose of 5-HT for nerve discharge was 13 micro g/kg. The time- and dose-dependent kinetics of the nerve discharge rate were similar to the change of heart rate. The afferent neuronal discharge was mimicked by a selective 5-HT3 receptor agonist, 1-phenylbiguanide hydrochloride (PBA), and inhibited by a selective 5-HT3 antagonist, granisetron. The 5-HT(3/4) agonist, cisapride partially activated the vagus nerve, but the 5-HT4 agonist, RS6733 had no effect on the vagal afferent activity. Intra-gastric perfusion of lidocaine, moreover, abolished the 5-HT-induced vagal activation. These results indicate that the 5-HT transmission signal in the gastric mucosa inputs to the brain stem via 5-HT3 receptor-mediated vagal nerve afferent.     

Capsaicin-sensitive vagal fibres and 5-HT3-, gastrin releasing peptide- and cholecystokinin A-receptors are involved in distension-induced inhibition of gastric emptying in the rat.

The present study was undertaken to investigate how the activation of gastric mechanoreceptors by distension of the stomach in conscious gastric fistula rats influences gastric emptying; and the roles of capsaicin sensitive vagal afferent fibres and the 5-HT3, GRP and CCK-A receptors involved in mediating these responses. To activate mechanoreceptors by non-nutrient dependent pathways, methylcellulose in saline was used to distend the stomach (5 cm H2O) and the subsequent emptying of saline was examined immediately, and at 3, 5 and 10 min following distension. Prior distension delayed the subsequent emptying of saline instilled into the stomach compared with non-distended controls (2.28+/-0.09 ml/5 min; P < 0.001). Topical application of capsaicin, completely abolished the distension-induced inhibition of gastric emptying when compared with vehicle treated rats (2.82+/-0.09 vs. 2.38+/-0.04 ml/5 min; P < 0.001). Peripheral administration of a GRP antagonist (2258 U89UJ, 1 mg/kg), and a 5-HT3 antagonist (BRL4369UA, 50 microg/kg) significantly reversed (2.56+/-0.14 ml/5 min; P < 0.05 and 2.61+/-0.07 ml/5 min; P < 0.01; respectively) the delay in gastric emptying induced by distension. When the rats were treated with the CCK-A antagonist, gastric emptying of saline following distension was also significantly facilitated (2.56+/-0.07 ml/5 min; P < 0.001). In contrast, the CCK-B/gastrin receptor antagonist had no significant effect on the distension induced delay in gastric emptying (1.95+/-0.12 ml/5 min). The present results suggest that gastric distension in conscious gastric fistula rats delays gastric emptying by activating capsaicin-sensitive extrinsic afferent nerve fibres. Moreover, the results also indicate that distension-induced mechanisms involve GRP, 5-HT3 and CCK-A receptors, but not CCK-B receptors.     

Gastric distension-induced release of 5-HT stimulates c-fos expression in specific brain nuclei via 5-HT3 receptors in conscious rats

We examined c-fos expression in specific brain nuclei in response to gastric distension and investigated whether 5-HT released from enterochromaffin (EC) cells was involved in this response. The role of 5-HT3 receptors in this mechanism was also addressed. Release of 5-HT was examined in an ex vivo-perfused stomach model, whereas c-fos expression in brain nuclei induced by gastric distension was examined in a freely moving conscious rat model. Physiological levels of gastric distension stimulated the vascular release of 5-HT more than luminal release of 5-HT, and induced c-fos expression in the nucleus of the solitary tract (NTS), area postrema (AP), paraventricular nucleus (PVN), and supraoptic nucleus (SON). The c-fos expression in all these brain nuclei was blocked by truncal vagotomy as well as by perivagal capsaicin treatment, suggesting that vagal afferent pathways may mediate this response. Intravenous injection of 5-HT3 receptor antagonist granisetron blocked c-fos expression in all brain nuclei examined, although intracerebroventricular injection of granisetron had no effect, suggesting that 5-HT released from the stomach may activate 5-HT3 receptors located in the peripheral vagal afferent nerve terminals and then induce brain c-fos expression. c-fos Positive cells in the NTS were labeled with retrograde tracer fluorogold injected in the PVN, suggesting that neurons in the NTS activated by gastric distension project axons to the PVN. The present results suggest that gastric distension stimulates 5-HT release from the EC cells and the released 5-HT may activate 5-HT3 receptors located on the vagal afferent nerve terminals in the gastric wall leading to neuron activation in the NTS and AP and subsequent activation of neurons in the PVN and SON.     

Intestinal serotonin acts as paracrine substance to mediate pancreatic secretion stimulated by luminal factors

We recently demonstrated that luminal factors such as osmolality, disaccharides, and mechanical stimulation evoke pancreatic secretion by activating 5-hydroxytryptamine subtype 3 (serotonin-3, 5-HT3) receptors on mucosal vagal afferent fibers in the intestine. We hypothesized that 5-HT released by luminal stimuli acts as a paracrine substance, activating the mucosal vagal afferent fibers to stimulate pancreatic secretion. In the in vivo rat model, luminal perfusion of maltose or hypertonic NaCl increased 5-HT level threefold in intestinal effluent perfusates. Similar levels were observed after intraluminal 10(-5) M 5-HT perfusion. These treatments did not affect 5-HT blood levels. In a separate study, intraduodenal, but not intraileal, 5-HT application induced a dose-dependent increase in pancreatic protein secretion, which was not blocked by the CCK-A antagonist CR-1409. Acute vagotomy, methscopolamine, or perivagal or intestinal mucosal application of capsaicin abolished 5-HT-induced pancreatic secretion. In conscious rats, luminal 10(-5) M 5-HT administration produced a 90% increase in pancreatic protein output, which was markedly inhibited by the 5-HT3 antagonist ondansetron. In conclusion, luminal stimuli induce 5-HT release, which in turn activates 5-HT3 receptors on mucosal vagal afferent terminals. In this manner, 5-HT acts as a paracrine substance to stimulate pancreatic secretion via a vagal cholinergic pathway.     

Water stimulation of the posterior oral cavity induces inhibition of gastric motility

The response of gastric motility to the administration of water and saline in the larynx and epiglottis was investigated in urethan-chloralose anesthetized rats. Administration of water inhibited motility of the distal stomach, but 0.15 M NaCl did not induce the inhibitory response. Bilateral sectioning of the superior laryngeal nerve (SLN) abolished the inhibitory response induced by water. Bilateral cervical vagotomies abolished the inhibitory responses, although spinal transection did not affect the inhibitory response. These inhibitory responses have been observed in immobilized animals. The degree of inhibition by water and hypotonic saline was negatively correlated with the sodium concentration. In contrast, the degree of inhibition to hypertonic saline was positively correlated with the sodium concentration. The proximal stomach also showed a reduction in intragastric pressure in response to the administration of water. These findings suggest that water-responsive afferent neurons in the SLN suppress gastric motility via the vagal efferent nerve.     

Spatial organization of neurons in the dorsal motor nucleus of the vagus synapsing with intragastric cholinergic and nitric oxide/VIP neurons in the rat

The dorsal motor nucleus of the vagus (DMV) contains preganglionic neurons that control gastric motility and secretion. Stimulation of different parts of the DMV results in a decrease or an increase in gastric motor activities, suggesting a spatial organization of vagal preganglionic neurons in the DMV. Little is known about how these preganglionic neurons in the DMV synapse with different groups of intragastric motor neurons to mediate contraction or relaxation of the stomach. We used pharmacological and immunohistochemical methods to characterize intragastric neural pathways involved in mediating gastric contraction and relaxation in rats. Microinjections of L-glutamate (L-Glu) into the rostral or caudal DMV produced gastric contraction and relaxation, respectively, in a dose-related manner. Intravenous infusion of hexamethonium blocked these actions, suggesting mediation via preganglionic cholinergic pathways. Atropine inhibited gastric contraction by 85.5 +/- 4.5%. Gastric relaxation was reduced by intravenous administration of N(G)-nitro-L-arginine methyl ester (L-NAME; 52.5 +/- 11.9%) or VIP antagonist (56.3 +/- 14.9%). Combined administration of L-NAME and VIP antagonist inhibited gastric relaxation evoked by L-Glu (87.8 +/- 4.3%). Immunohistochemical studies demonstrated choline acetyltransferase immunoreactivity in response to L-Glu microinjection into the rostral DMV in 88% of c-Fos-positive intragastric myenteric neurons. Microinjection of L-Glu into the caudal DMV evoked expression of nitric oxide (NO) synthase and VIP immunoreactivity in 81 and 39%, respectively, of all c-Fos-positive intragastric myenteric neurons. These data indicate spatial organization of the DMV. Depending on the location, microinjection of L-Glu into the DMV may stimulate intragastric myenteric cholinergic neurons or NO/VIP neurons to mediate gastric contraction and relaxation.     

Peripheral versus central modulation of gastric vagal pathways by metabotropic glutamate receptor 5

Metabotropic glutamate receptors (mGluR) are classified into group I, II, and III mGluR. Group I (mGluR1, mGluR5) are excitatory, whereas group II and III are inhibitory. mGluR5 antagonism potently reduces triggering of transient lower esophageal sphincter relaxations and gastroesophageal reflux. Transient lower esophageal sphincter relaxations are mediated via a vagal pathway and initiated by distension of the proximal stomach. Here, we determined the site of action of mGluR5 in gastric vagal pathways by investigating peripheral responses of ferret gastroesophageal vagal afferents to graded mechanical stimuli in vitro and central responses of nucleus tractus solitarius (NTS) neurons with gastric input in vivo in the presence or absence of the mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP). mGluR5 were also identified immunohistochemically in the nodose ganglia and NTS after extrinsic vagal inputs had been traced from the proximal stomach. Gastroesophageal vagal afferents were classified as mucosal, tension, or tension-mucosal (TM) receptors. MPEP (1-10 microM) inhibited responses to circumferential tension of tension and TM receptors. Responses to mucosal stroking of mucosal and TM receptors were unaffected. MPEP (0.001-10 nmol icv) had no major effect on the majority of NTS neurons excited by gastric distension or on NTS neurons inhibited by distension. mGluR5 labeling was abundant in gastric vagal afferent neurons and sparse in fibers within NTS vagal subnuclei. We conclude that mGluR5 play a prominent role at gastroesophageal vagal afferent endings but a minor role in central gastric vagal pathways. Peripheral mGluR5 may prove a suitable target for reducing mechanosensory input from the periphery, for therapeutic benefit.     

Glucose acts in the CNS to regulate gastric motility during hypoglycemia

Our purposes were to 1) develop an animal model where intravenously (iv) administered d-glucose consistently inhibited antral motility, and 2) use this model to assess whether iv glucose acts to inhibit motility from a peripheral or a central nervous system site and to elucidate the factor(s) that determine(s) whether stomach motor function is sensitive to changes in blood glucose. Rats were anesthetized with alpha-chloralose-urethane, and antral motility was measured by a strain-gauge force transducer sutured to the antrum. In some cases, antral motility and gastric tone were measured by monitoring intragastric balloon pressure. Increases in blood glucose were produced by continuous iv infusion of 25% d-glucose at 2 ml/h. Inhibition of antral motility and gastric tone was observed when gastric contractions were induced by hypoglycemia (subcutaneously administered insulin, 2.5 IU/animal). In contrast, no inhibition of gastric motor function was observed when glucose infusion was tested on gastric contractions that were 1) spontaneously occurring, 2) evoked by iv administered bethanechol in vagotomized animals, and 3) evoked by the TRH analog RX77368, microinjected into the dorsal motor nucleus of the vagus. Using the model of insulin-induced hypoglycemia to increase gastric motor activity, we found that neither sectioning the hepatic branch of the vagus (n = 5), nor treating animals with capsaicin to destroy sensory vagal afferent nerves (n = 5) affected the ability of iv d-glucose to inhibit gastric motor function. Our results indicate that an important factor determining whether stomach motor function will be sensitive to changes in blood glucose is the method used to stimulate gastric contractions, and that the primary site of the inhibitory action of iv glucose on gastric motility is the central nervous system rather than the periphery.     

Lack of interaction between peripheral injection of CCK and obestatin in the regulation of gastric satiety signaling in rodents

Obestatin is a new peptide for which anorexigenic effects were recently reported in mice. We investigate whether peripheral injection of obestatin or co-injection with cholecystokinin (CCK) can modulate food intake, gastric motor function (intragastric pressure and emptying) and gastric vagal afferent activity in rodents. Obestatin (30, 100 and 300 microg/kg, i.p.) did not influence cumulative food intake for the 2h post-injection in rats or mice nor gastric emptying in rats. In rats, obestatin (300 microg/kg) did not modify CCK (1 microg/kg, i.p.)-induced significant decrease in food intake (36.6%) and gastric emptying (31.0%). Furthermore, while rats injected with CCK (0.3 microg/kg, i.v.) displayed gastric relaxation, no change in gastric intraluminal pressure was elicited by obestatin (300 microg/kg, i.v.) pre- or post-CCK administration. In in vitro rat gastric vagal afferent preparations, 20 units that had non-significant changes in basal activity after obestatin at 30 microg responded to CCK at 10 ng by a 182% increase. These data show that obestatin neither influences cumulative food intake, gastric motility or vagal afferent activity nor CCK-induced satiety signaling.     

Secretin activates vagal primary afferent neurons in the rat: evidence from electrophysiological and immunohistochemical studies

In this study, we evaluated the vagal afferent response to secretin at physiological concentrations and localized the site of secretin's action on vagal afferent pathways in the rat. The discharge of sensory neurons supplying the gastrointestinal tract was recorded from nodose ganglia. Of 91 neurons activated by electrical vagal stimulation, 19 neurons showed an increase in firing rate in response to intestinal perfusion of 5-HT (from 1.5 +/- 0.2 to 25 +/- 4 impulses/20 s) but no response to intestinal distension. A close intra-arterial injection of secretin (2.5 and 5.0 pmol) elicited responses in 15 of these 19 neurons (from 1.5 +/- 0.2 impulses/20 s at basal to 21 +/- 4 and 43 +/- 5 impulses/20 s, respectively). Subdiaphragmatic vagotomy and perivagal application of capsaicin, but not supranodose vagotomy, completely abolished the secretin-elicited vagal nodose neuronal response. In a separate study, 9 tension receptor afferents among 91 neurons responded positively to intestinal distension but failed to respond to luminal 5-HT. These nine neurons also showed no response to administration of secretin. As expected, immunohistochemical studies showed that secretin administration significantly increased the number of Fos-positive neurons in vagal nodose ganglia. In conclusion, we demonstrated for the first time that vagal sensory neurons are activated by secretin at physiological concentrations. A subpopulation of secretin-sensitive vagal afferent fibers is located in the intestinal mucosa, many of which are responsive to luminal 5-HT.     

D-glucose modulates synaptic transmission from the central terminals of vagal afferent fibers

Experimental evidence suggests that glucose modulates gastric functions via vagally mediated effects. It is unclear whether glucose affects only peripheral vagal nerve activity or whether glucose also modulates vagal circuitry at the level of the brain stem. This study used whole cell patch-clamp recordings from neurons of the nucleus of the tractus solitarius (NTS) to assess whether acute variations in glucose modulates vagal brain stem neurocircuitry. Increasing D-glucose concentration induced a postsynaptic response in 40% of neurons; neither the response type (inward vs. outward current) nor response magnitude was altered in the presence of tetrodotoxin suggesting direct effects on the NTS neuronal membrane. In contrast, reducing d-glucose concentration induced a postsynaptic response (inward or outward current) in 54% of NTS neurons; tetrodotoxin abolished these responses, suggesting indirect sites of action. The frequency, but not amplitude, of spontaneous and miniature excitatory postsynaptic currents (EPSCs) was correlated with d-glucose concentration in 79% of neurons tested (n = 48). Prior surgical afferent rhizotomy abolished the ability of D-glucose to modulate spontaneous EPSC frequency, suggesting presynaptic actions at vagal afferent nerve terminals to modulate glutamatergic synaptic transmission. In experiments in which EPSCs were evoked via electrical stimulation of the tractus solitarius, EPSC amplitude correlated with D-glucose concentration. These effects were not mimicked by L-glucose, suggesting the involvement of glucose metabolism, not uptake, in the nerve terminal. These data suggest that the synaptic connections between vagal afferent nerve terminals and NTS neurons are a strong candidate for consideration as one of the sites where glucose-evoked changes in vagovagal reflexes occurs.     

Luminal amino acid sensing in the rat gastric mucosa

Recent advancements in molecular biology in the field of taste perception in the oral cavity have raised the possibility for ingested nutrients to be "tasted" in the upper gastrointestinal tract. The purpose of this study was to identify the existence of a nutrient-sensing system by the vagus in the rat stomach. Afferent fibers of the gastric branch increased their firing rate solely with the intragastric application of the amino acid glutamate. Other amino acids failed to have the same effect. This response to glutamate was blocked by the depletion of serotonin (5-HT) and inhibition of serotonin receptor(3) (5-HT(3)) or nitric oxide (NO) synthase enzyme. Luminal perfusion with the local anesthesia lidocaine abolished the glutamate-evoked afferent activation. The afferent response was also mimicked by luminal perfusion with a NO donor, sodium nitroprusside. In addition, the NO donor-induced afferent activation was abolished by 5-HT(3) blockade as well. Altogether, these results strongly suggest the existence of a sensing system for glutamate in the rat gastric mucosa. Thus luminal glutamate would enhance the electrophysiological firing rate of afferent fibers from the vagus nerve of the stomach through the production of mucosal bioactive substances such as NO and 5-HT. Assuming there is a universal coexistence of free glutamate with dietary protein, a glutamate-sensing system in the stomach could contribute to the gastric phase of protein digestion.     

Apolipoprotein A-IV stimulates duodenal vagal afferent activity to inhibit gastric motility via a CCK1 pathway

Apolipoprotein A-IV (apo A-IV), a peptide expressed by enterocytes in the mammalian small intestine and released in response to long-chain triglyceride absorption, may be involved in the regulation of gastric acid secretion and gastric motility. The specific aim of the present study was to determine the pathway involved in mediating inhibition of gastric motility produced by apo A-IV. Gastric motility was measured manometrically in response to injections of either recombinant purified apo A-IV (200 microg) or apo A-I, the structurally similar intestinal apolipoprotein not regulated by triglyceride absorption, close to the upper gastrointestinal tract in urethane-anesthetized rats. Injection of apo A-IV significantly inhibited gastric motility compared with apo A-I or vehicle injections. The response to exogenous apo A-IV injections was significantly reduced by 77 and 55%, respectively, in rats treated with the CCK(1) receptor blocker devazepide or after functional vagal deafferentation by perineural capsaicin treatment. In electrophysiological experiments, isolated proximal duodenal vagal afferent fibers were recorded in vitro in response to close-arterial injection of vehicle, apo A-IV (200 microg), or CCK (10 pmol). Apo A-IV stimulated the discharge of duodenal vagal afferent fibers, significantly increasing the discharge in 4/7 CCK-responsive units, and the response was abolished by CCK(1) receptor blockade with devazepide. These data suggest that apo A-IV released from the intestinal mucosa during lipid absorption stimulates the release of endogenous CCK that activates CCK(1) receptors on vagal afferent nerve terminals initiating feedback inhibition of gastric motility.     

Airway smooth muscle relaxation induced by 5-HT(2A) receptors: role of Na(+)/K(+)-ATPase pump and Ca(2+)-activated K(+) channels

AIMS: Although 5-hydroxytryptamine (5-HT) contracts airway smooth muscle in many mammalian species, in guinea pig and human airways 5-HT causes a contraction followed by relaxation. This study explored potential mechanisms involved in the relaxation induced by 5-HT. MAIN METHODS: Using organ baths, patch clamp, and intracellular Ca(2+) measurement techniques, the effect of 5-HT on guinea pig airway smooth muscle was studied. KEY FINDINGS: A wide range of 5-HT concentrations caused a biphasic response of tracheal rings. Response to 32 muM 5-HT was notably reduced by either tropisetron or methiothepin, and almost abolished by their combination. Incubation with 10 nM ketanserin significantly prevented the relaxing phase. Likewise, incubation with 100 nM charybdotoxin or 320 nM iberiotoxin and at less extent with 10 muM ouabain caused a significant reduction of the relaxing phase induced by 5-HT. Propranolol, L-NAME and 5-HT(1A), 5-HT(1B)/5-HT(1D) and 5-HT(2B) receptors antagonist did not modify this relaxation. Tracheas from sensitized animals displayed reduced relaxation as compared with controls. In tracheas precontracted with histamine, a concentration response curve to 5-HT (32, 100 and 320 muM) induced relaxation and this effect was abolished by charybdotoxin, iberiotoxin or ketanserin. In single myocytes, 5-HT in the presence of 3 mM 4-AP notably increased the K(+) currents (I(K(Ca))), and they were completely abolished by charybdotoxin, iberiotoxin or ketanserin. SIGNIFICANCE: During the relaxation induced by 5-HT two major mechanisms seem to be involved: stimulation of the Na(+)/K(+)-ATPase pump, and increasing activity of the high-conductance Ca(2+)-activated K(+) channels, probably via 5-HT(2A) receptors.