[99mTc]Tc-DGA1, a Promising CCK2R-Antagonist-Based Tracer for Tumor Diagnosis with Single-Photon Emission Computed Tomography.

Radiolabeled gastrin analogues have been proposed for theranostics of cholecystokinin subtype 2 receptor (CCK2R)-positive cancer. Peptide radioligands based on other receptor antagonists have displayed superior pharmacokinetics and higher biosafety than agonists. Here, we present DGA1, a derivative of the nonpeptidic CCK2R antagonist Z-360 carrying an acyclic tetraamine, for [99mTc]Tc labeling. Preclinical comparison of [99mTc]Tc-DGA1 with [99mTc]Tc-DG2 (CCK2R-agonist reference) was conducted in HEK293-CCK2R/CCK2i4svR cells and mice models, qualifying [99mTc]Tc-DGA1 for further study in patients with CCK2R-positive tumors and single-photon emission computed tomography/CT.

Heterogeneous effects of cholecystokinin on neuronal response properties in deep layers of rat barrel cortex.

Cholecystokinin (CCK) is one of the most studied neuropeptides in the brain. In this study, we investigated the effects of CCK-8s and LY225910 (CCK2 receptor antagonist) on properties of neuronal response to natural stimuli (whisker deflection) in deep layers of rat barrel cortex. This study was done on 20 male Wistar rats, weighing 230-260 g. CCK-8s (300 nmol/rat) and LY225910 (1 µmol/rat) were administered intracerebroventricularly (ICV). Neuronal responses to deflection of principal (PW) and adjacent (AW) whiskers were recorded in the barrel cortex using tungsten microelectrodes. Computer controlled mechanical displacement was used to deflect whiskers individually or in combination at 30 ms inter-stimulus intervals. ON and OFF responses for PW and AW deflections were measured. A condition-test ratio (CTR) was computed to quantify neuronal responses to whisker interaction. ICV administration of CCK-8s and LY225910 had heterogeneous effects on neuronal spontaneous activity, ON and OFF responses to PW and/or AW deflections, and CTR for both ON and OFF responses. The results of this study demonstrated that CCK-8s can modulate neuronal response properties in deep layers of rat barrel cortex probably via CCK2 receptors.

CCK reduces the food intake mainly through CCK1R in Siberian sturgeon (Acipenser baerii Brandt).

To explore the effect of CCK on food intake in Siberian sturgeon, cck cDNA sequence of 1005 bp was obtained, and cck mRNA possessed the highest expression in brain. The expressions of cck were significantly increased after feeding 1 and 3 h, while displaying significant decrease after fasting within 15 days in brain and duodenum. Re-feeding for 3 days induced cck level returned to basic level. Acute i.p. injection experiment showed 100 and 200 ng/g BW CCK8 inhibited the food intake in 0-1 h together with the cumulative food intake within 3 h. 7 days chronic i.p. injection of 100 and 200 ng/g BW CCK8, both daily food intake and cumulative food intake were significantly decreased. In addition, chronic i.p injection of CCK8 induced the expression of feeding related factors changes including cck, ucn3, cart, apelin, pyy and npy in respective organization. Moreover, as revealed by the results, Lorglumide, the CCK1R selective antagonist, effectively reversed the inhibitory effects of CCK8 on food intake and the levels of feeding related factors. On the other hand, LY 225910, the CCK2R selective antagonist, partially reversed these effects. These results indicate CCK is a satiety factor inhibits the feeding of Siberian sturgeon primarily through CCK1R.

Intrarenal signaling mediated by CCK plays a role in salt intake-induced natriuresis.

The natriuretic hormone CCK exhibits its gene transcripts in total kidney extracts. To test the possibility of CCK acting as an intrarenal mediator of sodium excretion, we examined mouse kidneys by 1) an in situ hybridization technique for CCK mRNA in animals fed a normal- or a high-sodium diet; 2) immuno-electron microscopy for the CCK peptide, 3) an in situ hybridization method and immunohistochemistry for the CCK-specific receptor CCKAR; 4) confocal image analysis of receptor-mediated Ca2+ responses in isolated renal tubules; and 5) metabolic cage experiments for the measurement of urinary sodium excretion in high-salt-fed mice either treated or untreated with the CCKAR antagonist lorglumide. Results showed the CCK gene to be expressed intensely in the inner medulla and moderately in the inner stripe of the outer medulla, with the expression in the latter being enhanced by high sodium intake. Immunoreactivity for the CCK peptide was localized to the rough endoplasmic reticulum of the medullary interstitial cells in corresponding renal regions, confirming it to be a secretory protein. Gene transcripts, protein products, and the functional activity for CCKAR were consistently localized to the late proximal tubule segments (S2 and S3) in the medullary rays, and the outer stripe of the outer medulla. Lorglumide significantly diminished natriuretic responses of mice to a dietary sodium load without altering the glomerular filtration rate. These findings suggest that the medullary interstitial cells respond to body fluid expansion by CCK release for feedback regulation of the late proximal tubular reabsorption.

Potential roles for calcium-sensing receptor (CaSR) and transient receptor potential ankyrin-1 (TRPA1) in murine anorectic response to deoxynivalenol (vomitoxin).

Food contamination by the trichothecene mycotoxin deoxynivalenol (DON, vomitoxin) has the potential to adversely affect animal and human health by suppressing food intake and impairing growth. In mice, the DON-induced anorectic response results from aberrant satiety hormone secretion by enteroendocrine cells (EECs) of the gastrointestinal tract. Recent in vitro studies in the murine STC-1 EEC model have linked DON-induced satiety hormone secretion to activation of calcium-sensing receptor (CaSR), a G-coupled protein receptor, and transient receptor potential ankyrin-1 (TRPA1), a TRP channel. However, it is unknown whether similar mechanisms mediate DON's anorectic effects in vivo. Here, we tested the hypothesis that DON-induced food refusal and satiety hormone release in the mouse are linked to activation of CaSR and TRPA1. Oral treatment with selective agonists for CaSR (R-568) or TRPA1 (allyl isothiocyanate (AITC)) suppressed food intake in mice, and the agonist's effects were suppressed by pretreatment with corresponding antagonists NPS-2143 or ruthenium red (RR), respectively. Importantly, NPS-2143 or RR inhibited both DON-induced food refusal and plasma elevations of the satiety hormones cholecystokinin (CCK) and peptide YY3-36 (PYY3-36); cotreatment with both antagonists additively suppressed both anorectic and hormone responses to DON. Taken together, these in vivo data along with prior in vitro findings support the contention that activation of CaSR and TRPA1 contributes to DON-induced food refusal by mediating satiety hormone exocytosis from EEC.

The Nocebo Effect.

A growing body of evidence is emerging for a phenomenon known as the nocebo effect. This is when a person is conditioned to expect a negative response, or to anticipate negative effects from an experience. These findings highlight the importantance of effective communication with patients and the influence that good anxiety and pain management control can have in improving treatment outcomes. The placebo effect has been widely researched, but new studies have shown that nocebo can have a greater effect than placebo The nocebo effect is prevalent in interactions between patients and healthcare workers. Research has demonstrated that if a patient deems a healthcare professional not to understand or believe them, this can cause distress, and the physiological effect can reduce the prognosis of treatment. It has also been demonstrated that patients who are anxious or expect pain during a procedure, feel more pain because of this negative expectation.

Involvement of myristoylated alanine-rich C kinase substrate phosphorylation and translocation in cholecystokinin-induced amylase release in rat pancreatic acini.

Cholecystokinin (CCK) is a gastrointestinal hormone that induces exocytotic amylase release in pancreatic acinar cells. The activation of protein kinase C (PKC) is involved in the CCK-induced pancreatic amylase release. Myristoylated alanine-rich C kinase substrate (MARCKS) is a ubiquitously expressed substrate of PKC. MARCKS has been implicated in membrane trafficking in several cell types. The phosphorylation of MARCKS by PKC results in the translocation of MARCKS from the membrane to the cytosol. Here, we studied the involvement of MARCKS in the CCK-induced amylase release in rat pancreatic acini. Employing Western blotting, we detected MARCKS protein in the rat pancreatic acini. CCK induced MARCKS phosphorylation. A PKC-δ inhibitor, rottlerin, inhibited the CCK-induced MARCKS phosphorylation and amylase release. In the translocation assay, we also observed CCK-induced PKC-δ activation. An immunohistochemistry study showed that CCK induced MARCKS translocation from the membrane to the cytosol. When acini were lysed by a detergent, Triton X-100, CCK partially induced displacement of the MARCKS from the GM1a-rich detergent-resistant membrane fractions (DRMs) in which Syntaxin2 is distributed. A MARCKS-related peptide inhibited the CCK-induced amylase release. These findings suggest that MARCKS phosphorylation by PKC-δ and then MARCKS translocation from the GM1a-rich DRMs to the cytosol are involved in the CCK-induced amylase release in pancreatic acinar cells.

EGFR Transactivation by Peptide G Protein-Coupled Receptors in Cancer.

Lung cancer kills approximately 1.3 million citizens in the world annually. The tyrosine kinase inhibitors (TKI) erlotinib and gefitinib are effective anti-tumor agents especially in lung cancer patients with epidermal growth factor receptor (EGFR) mutations. The goal is to increase the potency of TKI in lung cancer patients with wild type EGFR. G protein-coupled receptors (GPCR) transactivate the wild type EGFR in lung cancer cells. The GPCR can be activated by peptide agonists causing phosphatidylinositol turnover or stimulation of adenylylcyclase. Recently, nonpeptide antagonists were found to inhibit the EGFR transactivation caused by peptides. Nonpeptide antagonists for bombesin (BB), neurotensin (NTS) and cholecystokinin (CCK) inhibit lung cancer growth and increase the cytotoxicity of gefitinib. The results suggest that GPCR transactivation of the EGFR may play an important role in cancer cell proliferation.

Intraperitoneal CCK and fourth-intraventricular Apo AIV require both peripheral and NTS CCK1R to reduce food intake in male rats.

Apolipoprotein AIV (Apo AIV) and cholecystokinin (CCK) are secreted in response to fat consumption, and both cause satiation via CCK 1 receptor (CCK-1R)-containing vagal afferent nerves to the nucleus of the solitary tract (NTS), where Apo AIV is also synthesized. Fasted male Long-Evans rats received ip CCK-8 or fourth-ventricular (i4vt) Apo AIV alone or in combination. Food intake and c-Fos proteins (a product of the c-Fos immediate-early gene) were assessed. i4vt Apo AIV and/or ip CCK at effective doses reduced food intake and activated c-Fos proteins in the NTS and hypothalamic arcuate nucleus and paraventricular nucleus. Blockade of the CCK-1R by i4vt lorglumide adjacent to the NTS attenuated the satiating and c-Fos-stimulating effects of CCK and Apo AIV, alone or in combination. Maintenance on a high-fat diet (HFD) for 10 weeks resulted in weight gain and attenuation of both the behavioral and c-Fos responses to a greater extent than occurred in low-fat diet-fed and pair-fed HFD animals. These observations suggest that NTS Apo AIV or/and peripheral CCK requires vagal CCK-1R signaling to elicit satiation and that maintenance on a HFD reduces the satiating capacity of these 2 signals.

Regulation of aggression by obesity-linked genes TfAP-2 and Twz through octopamine signaling in Drosophila.

In Drosophila, the monoamine octopamine, through mechanisms that are not completely understood, regulates both aggression and mating behavior. Interestingly, our study demonstrates that the Drosophila obesity-linked homologs Transcription factor AP-2 (TfAP-2; TFAP2B in humans) and Tiwaz (Twz; KCTD15 in humans) interact to modify male behavior by controlling the expression of Tyramine ß-hydroxylase and Vesicular monanime transporter, genes necessary for octopamine production and secretion. Furthermore, we reveal that octopamine in turn regulates aggression through the Drosophila cholecystokinin satiation hormone homolog Drosulfakinin (Dsk). Finally, we establish that TfAP-2 is expressed in octopaminergic neurons known to control aggressive behavior and that TfAP-2 requires functional Twz for its activity. We conclude that genetically manipulating the obesity-linked homologs TfAP-2 and Twz is sufficient to affect octopamine signaling, which in turn modulates Drosophila male behavior through the regulation of the satiation hormone Dsk.

Simultaneous assessment of gastric emptying and secretion in rats by a novel computed tomography-based method.

Gastric emptying and gastric secretion are two major physiological functions of the stomach. The assessment of these functions in particular in small animals is challenging; no method currently available allows the simultaneous measurement of both functions, and methods used are lethal or invasive and often limited by spatial, temporal, or quantitative resolution. Here, we report the establishment and validation of a quantitative noninvasive high-throughput computed tomography-based method to measure simultaneously gastric emptying and secretion in rats in vivo. The imaging strategy enables one to visualize stomach anatomy and to quantify stomach volume and stomach contrast agent content. The method was validated by comparing the results to classical lethal methods (stomach phenol red content and stomach wet weight). Additionally, we showed that the use of a mild anesthetic does not interfere with normal gastric function, thereby enabling high-resolution temporal studies within single animals. These combined advantages were applied to reevaluate the impact of cholecystokinin (CCK), histamine, and oral glucose solutions on gastric function with high temporal resolution. CCK inhibited gastric emptying completely for 20 min, leading to the accumulation of gastric juice in the stomach. The CCK antagonist devazepide blocked this effect. Histamine stimulated both gastric secretion and delayed emptying. Oral glucose solution emptied at a fixed rate of 24-31 cal/min and stimulated gastric secretion. These results confirm previous observations and add volumetric changes as a new dimension. As computed tomography scanners become broadly available, this method is an excellent approach to measure the combined gastric functional readout and to reduce the number of animals used.

Reduced CCK signaling in obese-prone rats fed a high fat diet.

Deficits in satiation signaling during obesogenic feeding have been proposed to play a role in hyperphagia and weight gain in animals prone to become obese. However, whether this impaired signaling is due to high fat (HF) feeding or to their obese phenotype is still unknown. Therefore, in the current study, we examined the effects of CCK-8 (0.5, 1.0, 2.0, and 4.0 µg/kg) on suppression of food intake of HF-fed obese prone (OP) and resistant (OR) rats. Additionally, we determined the role of endogenous CCK in lipid-induced satiation by measuring plasma CCK levels following a lipid gavage, and tested the effect of pretreatment with devazepide, a CCK-1R antagonist on intragastric lipid-induced satiation. Finally, we examined CCK-1R mRNA levels in the nodose ganglia. We show that OP rats have reduced feeding responses to the low doses of exogenous CCK-8 compared to OR rats. Furthermore, OP rats exhibit deficits in endogenous CCK signaling, as pretreatment with devazepide failed to abolish the reduction in food intake following lipid gavage. These effects were associated with reduced plasma CCK after intragastric lipid in OP but not OR rats. Furthermore, HF feeding resulted in downregulation of CCK-1Rs in the nodose ganglia of OP rats. Collectively, these results demonstrate that HF feeding leads to impairments in lipid-induced CCK satiation signaling in obese-prone rats, potentially contributing to hyperphagia and weight gain.

Spinal transient receptor potential ankyrin 1 channel induces mechanical hypersensitivity, increases cutaneous blood flow, and mediates the pronociceptive action of dynorphin A.

We characterized pain behavior and cutaneous blood flow response induced by activation of the spinal transient receptor potential ankyrin 1 (TRPA1) channel using intrathecal drug administrations in the rat. Additionally, we assessed whether the pronociceptive actions induced by intrathecally administered dynorphin A, cholecystokinin or prostaglandin F(2α) are mediated by the spinal TRPA1 channel. Cinnamaldehyde, a TRPA1 agonist, produced a dose-related (3-10 µg) cutaneous blood flow increase and mechanical hypersensitivity effect. These effects at the currently used doses were of short duration and attenuated, although not completely, by pretreatment with A-967079, a TRPA1 antagonist. The cinnamaldehyde-induced hypersensitivity was also reduced by pretreatment with minocycline (an inhibitor of microglial activation), but not by carbenoxolone (a gap junction decoupler). In vitro study, however, indicated that minocycline only poorly blocks the TRPA1 channel. The mechanical hypersensitivity effect induced by dynorphin A, but not that by cholecystokinin or prostaglandin F(2α), was attenuated by a TRPA1 antagonist Chembridge-5861528 as well as A-967079. The cinnamaldehyde-induced cutaneous blood flow increase was not suppressed by MK-801, an NMDA receptor antagonist, or bicuculline, a GABA(A) receptor antagonist. The results indicate that spinal TRPA1 channels promote mechanical pain hypersensitivity and due to antidromic activation of nociceptive nerve fibers increase cutaneous blood flow. The attenuation of the cinnamaldehyde-induced hypersensitivity effect by minocycline may be explained by action other than block of the TRPA1 channel. Moreover, the spinal TRPA1 channel is involved in mediating the pronociceptive action of dynorphin A, but not that of the spinal cholecystokinin or prostaglandin F(2α).

Peptide YY3-36 and 5-hydroxytryptamine mediate emesis induction by trichothecene deoxynivalenol (vomitoxin).

Deoxynivalenol (DON, vomitoxin), a trichothecene mycotoxin produced by Fusarium sp. that frequently occurs in cereal grains, has been associated with human and animal food poisoning. Although a common hallmark of DON-induced toxicity is the rapid onset of emesis, the mechanisms for this adverse effect are not fully understood. Recently, our laboratory has demonstrated that the mink (Neovison vison) is a suitable small animal model for investigating trichothecene-induced emesis. The goal of this study was to use this model to determine the roles of two gut satiety hormones, peptide YY3-36 (PYY3-36) and cholecystokinin (CCK), and the neurotransmitter 5-hydroxytryptamine (5-HT) in DON-induced emesis. Following ip exposure to DON at 0.1 and 0.25mg/kg bw, emesis induction ensued within 15-30min and then persisted up to 120min. Plasma DON measurement revealed that this emesis period correlated with the rapid distribution and clearance of the toxin. Significant elevations in both plasma PYY3-36 (30-60min) and 5-HT (60min) but not CCK were observed during emesis. Pretreatment with the neuropeptide Y2 receptor antagonist JNJ-31020028 attenuated DON- and PYY-induced emesis, whereas the CCK1 receptor antagonist devezapide did not alter DON's emetic effects. The 5-HT3 receptor antagonist granisetron completely suppressed induction of vomiting by DON and the 5-HT inducer cisplatin. Granisetron pretreatment also partially blocked PYY3-36-induced emesis, suggesting a potential upstream role for this gut satiety hormone in 5-HT release. Taken together, the results suggest that both PYY3-36 and 5-HT play contributory roles in DON-induced emesis.

Human chorionic gonadotropin regulates gastric emptying in ovariectomized rats.

Prolongation of gastrointestinal transit resulting in nausea and vomiting in pregnancy (NVP) is the most common phenomenon during the first trimester of pregnancy. Increased human chorionic gonadotropin (hCG) concentration during the first trimester is the most likely cause of NVP. The aim of this study was to investigate the effect of hCG on gastrointestinal transit and plasma concentrations of cholecystokinin (CCK) in ovariectomized (Ovx) rats. I.p. injection of hCG was used to evaluate the dose effect of hCG on gastrointestinal transit in Ovx rats. The CCK antagonist lorglumide was used to clarify the role of CCK in regulating gastrointestinal transit. Gastrointestinal transit was assessed 15 min after intragastric gavage of a mixture of 10% charcoal and Na(2)(51)CrO(4) (0.5 µCi/ml). After i.p. administration of hCG, gastric emptying was inhibited in Ovx rats, but intestinal transit was not affected. Plasma CCK concentrations were increased in a dose-dependent manner after hCG treatment, and gastric emptying showed a significant negative correlation with CCK concentrations (P=0.01, r(2)=-0.5104). Peripheral administration (i.p.) of lorglumide, a selective CCK(1) receptor antagonist, attenuated the hCG-induced inhibition of gastric emptying in Ovx rats, whereas central administration via the i.c.v. route did not. hCG treatment of Ovx rats inhibits gastric emptying in a dose-dependent manner via a peripheral mechanism of CCK hypersecretion and activation of CCK(1) receptors.

Cardiovascular and inflammatory response to cholecystokinin during endotoxemic shock.

Cholecystokinin (CCK) was first described as a gastrointestinal hormone, but its receptors have been located in cardiac and vascular tissues, as well as in immune cells. Our aims were to investigate the role of CCK on lipopolysaccharide (LPS)-induced hypotension and its ability to modulate previously reported inflammatory mediators, therefore affecting cardiovascular function. To conduct these experiments, rats had their jugular vein cannulated for drug administration, and also, the femoral artery cannulated for mean arterial pressure (MAP) and heart rate records. Endotoxemia induced by LPS from Escherichia coli (1.5 mg/kg; i.v.) stimulated the release of CCK, a progressive drop in MAP, and increase in heart rate. Plasma tumor necrosis factor α (TNF-α), interleukin 10 (IL-10), nitrate, vasopressin, and lactate levels were elevated in the endotoxemic rats. The pretreatment with proglumide (nonselective CCK antagonist; 30 mg/kg; i.p.) aggravated the hypotension and also increased plasma TNF-α and lactate levels. On the other hand, CCK (0.4 µg/kg; i.v.) administered before LPS significantly restored MAP, reduced aortic and hepatic inducible nitric oxide synthase (iNOS) production, and elevated plasma vasopressin and IL-10 concentrations; it did not affect TNF-α. Physiological CCK concentration reduced nitrite and iNOS synthesis by peritoneal macrophages, possibly through a self-regulatory IL-10-dependent mechanism. Together, these data suggest a new role for the peptide CCK in modulating MAP, possibly controlling the inflammatory response, stimulating the anti-inflammatory cytokine, IL-10, and reducing vascular and macrophage iNOS-derived nitric oxide production. Based on these findings, CCK could be used as an adjuvant therapeutic agent to improve cardiovascular function.

TRPM5 is critical for linoleic acid-induced CCK secretion from the enteroendocrine cell line, STC-1.

Fatty acid-induced stimulation of enteroendocrine cells leads to release of the hormones such as cholecystokinin (CCK) that contribute to satiety. Recently, the fatty acid activated G protein-coupled receptor GPR120 has been shown to mediate long-chain unsaturated free fatty acid-induced CCK release from the enteroendocrine cell line, STC-1, yet the downstream signaling pathway remains unclear. Here we show that linoleic acid (LA) elicits membrane depolarization and an intracellular calcium rise in STC-1 cells and that these responses are significantly reduced when activity of G proteins or phospholipase C is blocked. LA leads to activation of monovalent cation-specific transient receptor potential channel type M5 (TRPM5) in STC-1 cells. LA-induced TRPM5 currents are significantly reduced when expression of TRPM5 or GPR120 is reduced using RNA interference. Furthermore, the LA-induced rise in intracellular calcium and CCK secretion is greatly diminished when expression of TRPM5 channels is reduced using RNA interference, consistent with a role of TRPM5 in LA-induced CCK secretion in STC-1 cells.

Placebo effect in patients with irritable bowel syndrome.

The placebo effect has evolved from being considered a nuisance factor in clinical research to a hot topic of scientific investigation. New research findings show that a placebo has real psychobiological and biological effects that are attributable to the overall therapeutic context. Irritable bowel syndrome (IBS) is a functional disorder of the gastrointestinal tract that shows a significant placebo response of around 40­50% among different clinical trials.A positive patient-practitioner relationship can enhance the placebo effect in IBS patients.Emerging literature using functional brain imaging has started to document the neuronal changes associated with the placebo phenomenon in IBS patients, showing aberrant neural network during visceral placebo analgesia when compared to controls. Further promotion and integration of laboratory and clinical research are encouraged to advance the understanding of placebo mechanisms in IBS patients.

Cholecystokinin facilitates neuronal excitability in the entorhinal cortex via activation of TRPC-like channels.

Cholecystokinin (CCK) is one of the most abundant neuropeptides in the brain, where it interacts with two G protein-coupled receptors (CCK-1 and CCK-2). Activation of both CCK receptors increases the activity of PLC, resulting in increases in intracellular calcium ion (Ca(2+)) release and activation of PKC. Whereas high density of CCK receptors has been detected in the superficial layers of the entorhinal cortex (EC), the functions of CCK in this brain region have not been determined. Here, we studied the effects of CCK on neuronal excitability of layer III pyramidal neurons in the EC. Our results showed that CCK remarkably increased the firing frequency of action potentials (APs). The effects of CCK on neuronal excitability were mediated via activation of CCK-2 receptors and required the functions of G proteins and PLC. However, CCK-mediated facilitation of neuronal excitability was independent of inositol trisphosphate receptors and PKC. CCK facilitated neuronal excitability by activating a cationic channel to generate membrane depolarization. The effects of CCK were suppressed by the generic, nonselective cationic channel blockers, 2-aminoethyldiphenyl borate and flufenamic acid, but potentiated by gadolinium ion and lanthanum ion at 100 µM. Depletion of extracellular Ca(2+) also counteracted CCK-induced increases in AC firing frequency. Moreover, CCK-induced enhancement of neuronal excitability was inhibited significantly by intracellular application of the antibody to transient receptor potential channel 5 (TRPC5), suggesting the involvement of TRPC5 channels. Our results provide a cellular and molecular mechanism to help explain the functions of CCK in vivo.

Apelin stimulation of duodenal bicarbonate secretion: feeding-dependent and mediated via apelin-induced release of enteric cholecystokinin.

AIMS: Apelin peptides are the endogenous ligand of the G protein-coupled receptor APJ. Proposed actions include involvement in control of cardiovascular functions, appetite and body metabolism. We have investigated the effects of apelin peptides on duodenal bicarbonate secretion in vivo and the release of cholecystokinin (CCK) from acutely isolated mucosal cells and the neuroendocrine cell line STC-1. METHODS: Lewis × Dark Agouti rats had free access to water and, unless fasted overnight, free access to food. A segment of proximal duodenum was cannulated in situ in anaesthetized animals. Mucosal bicarbonate secretion was titrated (pH stat) and apelin was administered to the duodenum by close intra-arterial infusion. Total RNA was extracted from mucosal specimens, reverse transcripted to cDNA and the expression of the APJ receptor measured by quantitative real-time PCR. Apelin-induced release of CCK was measured using (1) cells prepared from proximal small intestine and (2) STC-1 cells. RESULTS: Even the lowest dose of apelin-13 (6 pmol kg⁻¹ h⁻¹) caused a significant rise in bicarbonate secretion. Stimulation occurred only in continuously fed animals and even a 100-fold greater dose (600 pmol kg⁻¹ h⁻¹) of apelin was without effect in overnight food-deprived animals. Fasting also induced an eightfold decrease in the expression of APJ receptor mRNA. Apelin induced significant release of CCK from both mucosal and STC-1 cells, and the CCK(A) receptor antagonist devazepide abolished bicarbonate secretory responses to apelin. CONCLUSION: Apelin-induced stimulation of duodenal electrolyte secretion is feeding-dependent and mediated by local mucosal release of CCK.