CCK mRNA Levels Research Articles

Role of Cholecystokinin (cck) in Feeding Regulation of Largemouth Bass (Micropterus salmoides): Peptide Activation and Antagonist Inhibition.

This study investigated the role of cholecystokinin (cck) in the feeding regulation of largemouth bass (Micropterus salmoides) via peptide activation and antagonist inhibition. The results show that the cck gene was expressed in various tissues, with the highest expression level occurring in the brain. Feeding, continuous feeding, and refeeding after fasting could significantly improve the mRNA levels of cck in the brain. Moreover, the activation of cck via injecting an exogenous CCK peptide could inhibit feed intake by regulating the mRNA levels of anorexigenic and feed-promoting factors in the brain and intestine. Furthermore, the CCK peptide reduced feed intake; however, the presence of an antagonist (Ly225910-CCK1R and devazepide-CCK2R) could reverse this effect through regulating the mRNA levels of anorexigenic and feed-promoting factors in the brain and intestine. Treatment with devazepide + CCK (CCK2R) reversed feed intake more effectively than Ly225910 + CCK (CCK1R) treatment. In summary, cck could regulate the feed intake of largemouth bass through regulating feeding-related genes in the brain and intestine. In addition, cck required binding with the receptor to inhibit feed intake more effectively in largemouth bass, and the binding effect of CCK1R was better than that of CCK2R.

Effects of fasting and environmental factors on appetite regulators in pond loach Misgurnus anguillicaudatus

The pond loach (Misgurnus anguillicaudatus) is an important aquaculture freshwater species, used as an ornamental fish, food source for humans and angling bait. Pond loaches are resistant to fasting and extreme environmental conditions, including temperature and low oxygen levels. Little is known about how these factors affect the feeding physiology and the endocrine regulation of feeding of loaches. In this study, we examined the effects of fasting, as well as increased temperature and decreased oxygen levels on food intake and transcript levels of appetite regulators. Fasted fish had lower blood glucose levels, and lower expression levels of intestine CCK and PYY, and brain CART1, but had higher levels of brain orexin and ghrelin than fed fish. Fish held at 30 °C had higher food intake, glucose levels, and mRNA levels of intestine CCK and PYY, and brain CART2, but lower brain orexin levels than fish at 20 °C. Fish held at low oxygen levels had a lower food intake, higher intestine CCKa and ghrelin, and brain orexin, CART2 and ghrelin mRNA expression levels than fish held at high O2 levels. Our results suggest that fasting and high temperatures increase the expression of orexigenic and anorexigenic factors respectively, whereas the increase in expression of both orexigenic and anorexigenic factors in low O2 environments might not be related to their role in feeding, but possibly to protection from tissue damage. The results of our study might shed new light on how pond loaches are able to cope with extreme environmental conditions such as low food availability, extreme temperatures and hypoxia.

Cholecystokinin-mediated effect of insulin pathway on the steroidogenic activity of follicular granulosa cells in Camelus bactrianus: Invitro study.

The current study aimed to explore the molecular mechanism by which the cholecystokinin (CCK)-mediated CCKAR and CCKBR, as well as the molecular mechanisms of CCK-mediated insulin signalling pathway, regulate oestrogen in the granulosa cells. Also, the expression of CCK in ovaries, uterus, hypothalamus and pituitary gland was investigated in Camelus bactrianus. Ovaries, uterus, hypothalamus and pituitary gland were collected from six, three before ovulation (control) and three after ovulation, slaughtered Camelus bactrianus. Ovulation was induced by IM injection of seminal plasma before slaughtering in the ovulated group. The results showed that there were differences in the transcription and protein levels of CCK in various tissues before and after ovulation (p < .05, p < .01). After transfection with p-IRES2-EGFP-CCK, the mRNA and protein levels of CCK, CCKAR, CCKBR and ER in follicular granulosa cells were significantly upregulated (p < .05, p < .01), and the content of E2 was significantly upregulated (p < .01); On the contrary, after transfection with si-CCK, the mRNA and protein levels of CCK, CCKAR, CCKBR and ER in follicular granulosa cells were significantly downregulated (p < .05, p < .01), and the content of E2 was significantly downregulated (p < .01). Regulating CCK can affect the mRNA levels of INS, INSR, IGF and IGF-R. In summary, regulating the expression level of CCK can activate insulin-related signalling pathways by CCKR, thereby regulating the steroidogenic activity of granulosa cells.

Glucose mediates upregulation of Cck expression independent of cAMP signaling

Cholecystokinin (CCK) is a gut peptide hormone that is also produced by pancreatic β‐cells under conditions of metabolic stress and obesity. Treatment with exogenous CCK can alleviate obesity‐related diabetes in animal models without significant side effects. We have established that CCK is expressed primarily in the pancreatic β‐cell and that CCK is highly upregulated in ob/ob mice. We have also shown that elevated cellular cAMP stimulates β‐cell CCK expression and secretion. However, the specific islet cell types that express CCK and the regulation of Cck transcription in the islet has not been thoroughly explored. A more recent study from our lab showed that high glucose concentrations can upregulate Cck expression in mouse islets and human islets. We hypothesize that Cck expression is upregulated in response to various diabetogenic conditions and that CCK upregulation by glucose can occur independent of cAMP activation.Islets from 16‐week‐old C57BL/6J mice were harvested for use in a dose‐dependent 24‐hour study using different glucose concentrations: No glucose, 2.5mM, 7mM, 11mM &amp; 25mM. After 24‐hour treatment, islets were collected for RNA isolation and cDNA synthesis to identify changes in Cck mRNA expression after incubation with different glucose concentrations. Cck mRNA levels were elevated 4‐8 fold in islets treated with 25 mM glucose compared to islets treated with 0 to 11mM glucose. Notably, the hypoglycemic treatment of no glucose and 2.5mM showed a 4‐fold increase when compared to the 7mM and 11mM treatments. Following this experiment, we wanted to determine if high glucose would still increase Cck mRNA expression after inhibiting cAMP. Islets were pre‐incubated for 45 minutes with 10nM sulprostone, which activated prostaglandin E3 (EP3) receptor and inhibits adenylate cyclase. After inhibition of cAMP, we incubated the islets at 2.5mM and 25mM glucose concentrations like the prior experiment and observed changes in Cck mRNA expression. After treatment with sulprostone, we are still able to see a significant increase in Cck mRNA expression in islets treated with 25mM glucose compared to those treated with 2.5mM glucose, even with cAMP inhibition.This data suggests that there are alternative pathways that could lead to CCK upregulation in the pancreatic islet independent of cAMP. It also shows that hypoglycemia may also increase Cckexpression, which suggests that CCK is upregulated in response to several forms of metabolic stress. We can further explore the effectiveness of glucose to increase CCK expression by treating islets with cycloheximide and deoxyglucose to determine if glucose metabolism is required for upregulation of Cck.This information will help with our general understanding of how CCK functions in pancreatic islet and how Cck transcription is regulated through signaling pathways and transcriptional networks that regulate β‐cell function and identity under diabetogenic conditions.

Effects of fasting and re-feeding on the expression of CCK, PYY, hypothalamic neuropeptides, and IGF-related genes in layer and broiler chicks

Cholecystokinin (CCK) and peptide YY (PYY) have been investigated as gut hormones that send satiation signals to the brain in mammals. There is evidence that chicken PYY mRNA expression was the highest in the pancreas compared to other tissues. We recently suggested that insulin-like growth factor (IGF)-1 and its binding proteins (IGFBPs) may be involved in the appetite regulation system in chicks. In the present study, in order to evaluate the possible roles of CCK, PYY, and IGF-related proteins in the appetite regulation system in chicks, we analyzed changes in the mRNA levels of these genes in response to fasting and re-feeding in layer and hyperphagic broiler chicks. In layer chicks, 12 h of fasting reduced the mRNA levels of intestinal CCK, PYY, Y2 receptor, and pancreatic PYY, and these changes were reversed by 12 h of re-feeding. On the other hand, in broiler chicks 12 h of fasting reduced the mRNA levels of intestinal PYY and Y2 receptor, but not intestinal CCK and pancreatic PYY, and these changes were reversed by 12 h of re-feeding. Hypothalamic NPY mRNA significantly increased by 12 h of fasting in both chicks, and these changes were reversed by re-feeding. Also, 12 h of fasting significantly increased the mRNA levels of hypothalamic agouti-related protein and reduced the mRNA levels of hepatic IGF-1 only in broiler chicks, and 12 h of re-feeding did not change these. IGFBP-1 and -2 mRNA levels were markedly increased by 12 h of fasting in both chicks, and these changes were reversed by re-feeding. IGFBP-3 mRNA levels were increased by 12 h of fasting only in layer chicks, while re-feeding reduced the mRNA levels of IGFBP-3 in both types of chicks. These results suggest that several peripheral hormones, such as pancreatic PYY and intestinal CCK, may not play important roles in the regulation of food intake in broiler chicks.

Nutrients regulate gene expression levels of neuropeptide Y, melanin-concentrating hormone 1, and cholecystokinin in yellowtail (Seriola quinqueradiata)

Neuropeptide Y (NPY), melanin-concentrating hormone 1 (MCH1), and cholecystokinin (CCK) are appetite-regulatory hormones. To understand the regulation of npy, mch1, and cck mRNA expression by dietary nutrients, the following two experiments were performed in the marine, carnivorous yellowtail (Seriola quinqueradiata). Juvenile yellowtails were fed one of four different diets for one week in Experiment 1—high protein-based diets combined with either fat (HF diet) or carbohydrates and low protein-based diets combined with either fat (LF diet) or carbohydrates. After feeding, serum non-esterified fatty acid concentration increased in the HF and LF groups, and serum free amino acid concentration increased in all dietary groups. In post-feeding fish, serum glucose concentration increased only in the HF group. The highest npy and cck mRNA levels in the forebrain were observed in the HF group, among the four dietary groups before feeding. Furthermore, a significant decrease in the npy mRNA level in the forebrain, after feeding, was observed only in the HF group. A similar trend was observed for cck and mch1 levels in the forebrain. Pyloric cecal cck increased significantly after feeding, in the LC group. Following the intraperitoneal administration of glucose in Experiment 2, the npy and mch1 mRNA expression levels in the forebrain decreased significantly. These results suggest that NPY and MCH1 levels in yellowtail are mainly regulated by the serum glucose concentration derived from the feed, and they work as feeding stimulators.

Gene expression of nutrient-sensing molecules in I cells of CCK reporter male mice.

Cholecystokinin (CCK) is secreted from enteroendocrine I cells in response to fat, carbohydrate, and protein ingestion. Gene expression of nutrient-sensing molecules in I cells remains unclear, primarily due to the difficulty in distinguishing I cells from intestinal epithelial cells in vivo. In this study, we generated CCK reporter male mice in which the red fluorescence protein tdTomato (Tomato) is produced by activation of the native murine Cck promoter. Fluorescence microscopy revealed the presence of Tomato-positive cells in upper small intestine (SI), lower SI, and colon. Flow cytometer analysis revealed that Tomato-positive cells among epithelial cells of upper SI, lower SI, and colon occurred at the rate of 0.95, 0.54, and 0.06%, respectively. In upper SI and lower SI, expression levels of Cck mRNA were higher in Tomato-positive cells than those in Tomato-negative cells. The fatty acid receptors Gpr120, Gpr40, and Gpr43 and the oleoylethanolamide receptor Gpr119 were highly expressed in Tomato-positive cells isolated from SI, but were not found in Tomato-positive cells from colon. The glucose and fructose transporters Sglt1, Glut2, and Glut5 were expressed in both Tomato-positive cells and -negative cells, but these expression levels tended to be decreased in Tomato-positive cells from upper SI to colon. The peptide transporter Pept1 and receptor Gpr93 were expressed in both Tomato-positive cells and -negative cells, whereas Casr was expressed only in Tomato-positive cells isolated from SI. Thus, this transgenic mouse reveals that I cell number and gene expression in I cells vary according to region in the gastrointestinal tract.

Protective effect of cholecystokinin octapeptide on angiotensin II-induced apoptosis in H9c2 cardiomyoblast cells.

Cholecystokinin (CCK) and its receptors are expressed in mammalian cardiomyocytes and are involved in cardiovascular system regulation; however, the exact effect and underlying mechanism of CCK in cardiomyocyte apoptosis remain to be elucidated. We examined whether sulfated CCK octapeptide (CCK-8) protects H9c2 cardiomyoblast cells against angiotensin II (Ang II)-induced apoptosis. The H9c2 cardiomyoblasts were subjected to Ang II with or without CCK-8 and the viability and apoptotic rate were detected using a Cell Counting Kit-8 assay, Hoechst 33342 staining, terminal deoxyribonucleotide transferase-mediated nick-end labeling assays, and flow cytometry. In addition, specific antiapoptotic mechanisms of CCK-8 were investigated using specific CCK1 (Devazepide) or CCK2 (L365260) receptor antagonists, or the PI3K inhibitor LY294002. The expression of CCK, CCK1 receptor, CCK2 receptor, Akt, p-Akt, Bad, p-Bad, Bax, Bcl-2, and caspase-3 were detected by Western blot analysis and real-time polymerase chain reaction. We found that CCK and its receptor messenger RNA (mRNA) and protein are expressed in H9c2 cardiomyoblasts. Ang II-induced increased levels of CCK mRNA and protein expression and decreased levels of CCK1 receptor protein and mRNA. Pretreatment of CCK-8 attenuated Ang II-induced cell toxicity and apoptosis. In addition, pretreatment of H9c2 cells with CCK-8 markedly induced expression of p-Akt, p-bad, and Bcl-2 and decreased the expression levels of Bax and caspase-3. The protective effects of CCK-8 were partly abolished by Devazepide or LY294002. Our results suggest that CCK-8 protects H9c2 cardiomyoblasts from Ang II-induced apoptosis partly via activation of the CCK1 receptor and the phosphatidyqinositol-3 kinase/protein kinase B (PI3K/Akt) signaling pathway.

SUN-098 Goldfish Insulin: Molecular Cloning, Hepatic Regulation by Spexin, and Mechanisms for Feeding Control in Fish Model

Insulin is a key hormone for metabolism and glucose homeostasis. In mammals, it also acts as a satiety factor with inhibitory effect on food intake, but its biological actions in appetite control in lower vertebrates is still unknown. Spexin (SPX), a neuropeptide with pleotropic functions, has been recently confirmed to be a novel satiety factor in fish model via functional coupling with insulin, although the details of their interactions have yet to be elucidated. In this study, using goldfish as a model, the mechanism for feeding regulation by insulin and its functional interactions with SPX at hepatic level were examined. As a first step, goldfish insulin was cloned and found to be widely expressed at tissue level, especially with high levels of expression in the liver and visceral fat. In silico modeling also showed that the 3D structure of the mature peptide of goldfish insulin was highly comparable if not identical to its human counterpart. In goldfish liver cell culture, insulin mRNA level could be up-regulated by SPX treatment via PLC/IP3/PKC and Ca2+/CaM/CaMK-II pathways, while the opposite was true by removing endogenous SPX using immunoneutralization with SPX antiserum. In whole animal experiment, IP injection with insulin was found to inhibit feeding behavior and food consumption in goldfish. Similar treatment in vivo was also effective in elevating POMC, CART, CCK and leptin mRNA levels with concurrent drop in NPY, AGRP and apelin transcript expression in the telencephalon, optic tectum and hypothalamus, the brain areas known to be involved in feeding control in fish models. Using brain cell culture prepared from these three areas, similar changes in transcript expression for the respective orexigenic and anorexigenic factors were also noted with insulin treatment in a time- and dose-dependent manner. These findings, as a whole, suggest that SPX can act in an autocrine/paracrine manner to induce insulin expression in goldfish liver and the subsequence rise of insulin in circulation presumably can inhibit food intake through differential regulation of orexigenic/anorexigenic signals expressed in brain areas involved in appetite control in fish model.

Study on the regulation of brain–gut peptide by Shenling Baizhu San in functional diarrhea rats

Abstract Objective To explore the therapeutic mechanism of Shenling Baizhu San (SLBZS) on functional diarrhea (FDr) by studying the brain–gut axis and related neuropeptides. Methods Sixty male Wistar rats were randomly divided into the control group, model group, SLBZS-treated group and Montmorillonite Powder-treated group (MP-treated group) (n = 15/group). Rats received gavage after the establishment of functional diarrhea. An equal volume of SLBZS solution and Montmorillonite Powder (MP) solution was administered to the SLBZS-treated group and MP-treated group, respectively, and an equal volume of distilled water was administered to the control group and the model group. The chemical components and targets related to SLBZS were identified from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and Traditional Chinese Medicine Integrated Database (TCMID). The effective chemical components were screened based on oral bioavailability (OB) and drug like-index (DL), and their biological functions were analyzed by GlueGO. Based on this screening, the expression of Cholecystokinin (CCK) and Ghrelin in the hypothalamus of rats was detected by real-time PCR (RT-PCR) and western blotting. Results In this study, 72 effective components and 190 core targets of SLBZS were screened. SLBZS may regulate smooth muscle contraction, energy metabolism and other biological processes. The results of RT-PCR showed that in the model group, the expression of CCK mRNA (P = .001) and Ghrelin mRNA (P = .000) increased significantly. Compared with the model group, CCK mRNA (P = .007) and Ghrelin mRNA (P = .001) levels in SLBZS-treated rats were decreased significantly. The results of western blotting showed that in the model group, the protein expression of CCK (P = .001) and Ghrelin (P = .000) increased significantly. The protein levels of CCK (P = .001) and Ghrelin (P = .005) in the SLBZS-treated group were decreased significantly compared with the model group. Conclusion SLBZS improved functional diarrhea by regulating the brain–gut axis. Changes in the expressions of brain–gut peptide, CCK and Ghrelin might explain the pathogenesis of functional diarrhea related to brain–gut peptide and gastrointestinal hormone.

Cholecystokinin Expression in the Development of Postinfarction Heart Failure

Background/Aims: Cholecystokinin (CCK) is expressed in cardiomyocytes and may also play an important role in cardiovascular regulation. Clinical studies have shown that plasma CCK levels are an independent marker of cardiovascular mortality in cardiac disease. However, whether the development of postinfarction heart failure is associated with changes in CCK expression is unknown. Methods: To investigate CCK expression patterns and the association between CCK expression and heart functional parameters, we randomized male Sprague-Dawley rats into myocardial infarction (MI) or sham operation (SO) groups. CCK expression levels were assessed by western blotting, immunohistochemistry, real-time polymerase chain reaction (PCR), and enzyme-linked immunosorbent assay (ELISA) at different time points (2, 4 or 6 weeks) after surgery. Brain natriuretic peptide (BNP) concentrations were determined using Western blotting and ELISA, myocardial morphology was assessed by microscopy. Results: Plasma CCK and BNP levels were significantly increased in all the MI groups compared with the corresponding SO groups. However, the degree to which myocardial CCK mRNA and protein expression levels were increased the MI groups compared with the SO groups was greater in the 4- and 6-week groups than in the 2-week group. Furthermore, plasma CCK levels were positively correlated with BNP concentrations and left ventricular end-systolic diameter (LVDs) and significantly negatively correlated with the ejection fraction (EF) and shortening fraction (SF) in model animals. Conclusions: Heart failure progression after infarction is associated with upregulated CCK levels; thus, CCK may be useful as a novel marker of heart failure.

Molecular cloning, characterization, and expression analysis of ghrelin and cholecystokinin in the pigeon (Columba livia)

Ghrelin and cholecystokinin (CCK) are multifunctional peptides. In the current study, complete sequences of ghrelin (800 bp) and CCK (739 bp) were firstly cloned in Columba livia by using rapid amplification of cDNA ends (RACE) method. The open reading frames of ghrelin (351bp) and CCK (393bp) encoded 116 amino acids and 130 amino acids, respectively. Sequence comparison indicated that pigeon ghrelin and CCK shared high identity with those reported in other avian species. Quantitative real-time PCR analysis found that ghrelin and CCK mRNAs expressed in three intestinal segments of pigeon during development. Both ghrelin and CCK showed generally higher expressions at days posthatch than embryonic periods regardless of intestinal segments. In duodenum and ileum, the expressions of ghrelin and CCK mRNA reached the peak values at 8 d posthatch. Jejunum CCK mRNA level increased linearly after hatching, and reached the highest point at posthatch 28 d. Based on documented effects of long chain fatty acids (LCFAs) on pigeon ghrelin and CCK expression were also investigated in vitro. Higher concentrations (50 μM or 250 μM) of linoleic acid, α-linolenic acid or arachidonic acid can significantly increase ghrelin mRNA level in pigeon jejunum. However, for oleic acid, the induction of ghrelin gene expressions needed a lower concentration (5 μM). 5 μM of linoleic acid, α-linolenic acid or arachidonic acid and 250 μM palmitic acid repressed CCK expression significantly. A higher concentration (250 μM) of oleic acid or α-linolenic acid can up-regulate CCK mRNA level significantly. Our results indicated that ghrelin and CCK may act key functions in pigeon intestine development and their expressions could be regulated by LCFAs.

5-HT receptor-mediated modulation of granule cell inhibition after juvenile stress recovers after a second exposure to adult stress

Aversive experiences in early life are thought to dispose to psychopathologies such as mood or anxiety disorders. In a two-hit stress model, we assessed the effects of juvenile and/or adult stress on the 5-HT-mediated modulation of synaptic inhibition of ventral dentate gyrus granule cells. Combined but not single stress exposure led to a significant reduction in activity and increased anxiety-like behavior. Similarly, the 5-HT1A receptor-mediated inhibition of evoked inhibitory postsynaptic currents (IPSCs) of granule cells was only reduced in single stress exposed animals. This was also true for the number of granule cells responding with a 5-HT3 receptor-dependent burst of miniature IPSCs. 5-HT3 receptors are expressed on cholecystokinin (CCK)+ basket cells in the hippocampus. In fact, we observed a reduction of steady-state mRNA levels of CCK+ basket cell markers after single juvenile or adult stress and partial recovery after combined stress, thus matching the electrophysiological findings. Adaptive changes in 5-HT-mediated modulation of synaptic inhibition and CCK+ basket cells in the DG may help to maintain normal levels of anxiety after single juvenile or adult stress exposure, as indicated by the increased anxiety that accompanies the loss of this regulation upon combined stress.

Characterization of the endocrine, digestive and morphological adjustments of the intestine in response to food deprivation and torpor in cunner, Tautogolabrus adspersus

The cunner, Tautogolabrus adspersus, is a marine teleost endemic to the cold waters of the Northwest Atlantic Ocean. The cunner is non-migratory and is known for its remarkable ability to endure the freezing winter months with little to no food by entering a torpid/dormant state. To evaluate the physiological strategies employed by the cunner's intestinal tract to withstand food deprivation, fish were sampled for their gut after a four-week period of acute food deprivation during their summer (active/feeding) state, as well as after 4months of overwinter fasting. Digestive capacity was evaluated by measuring digestive enzyme activity and related mRNA transcript expression for trypsin, alkaline phosphatase, alanine aminopeptidase and lipase. In order to assess how gut hormones affect/are affected by acute fasting and torpor, we examined the intestinal mRNA expression of several putative appetite regulators, i.e. CCK, apelin, orexin and mTOR. Short-term summer fasting induced a reduction in the activity, but not the transcript expression, of all digestive enzymes examined as well as a reduction in gut apelin mRNA. Torpor induced a reduction in the activity of all enzymes with the exception of alanine aminopeptidase, and a decrease in mRNA levels of alanine aminopeptidase, orexin, CCK and mTOR. Our results suggest that both acute fasting and long-term fasting induce a reduction in the intestinal function of cunner, as evidenced by an overall decrease in the activities of digestive enzymes and mRNA expression of several factors involved in feeding and digestion.

Interaction between cholecystokinin and the fibroblast growth factor system in the ventral tegmental area of selectively bred high- and low-responder rats

Individual differences in the locomotor response to novelty have been linked to basal differences in dopaminergic neurotransmission. Mesolimbic dopaminergic outputs are regulated by cholecystokinin (CCK), a neuropeptide implicated in anxiety. In turn, CCK expression is regulated by fibroblast growth factor-2 (FGF2), which has recently been identified as an endogenous regulator of anxiety. FGF2 binds to the high-affinity fibroblast growth factor receptor-1 (FGF-R1) to regulate the development and maintenance of dopamine neurons in the ventral tegmental area (VTA). However, the relationship between the FGF and CCK systems in the VTA is not well understood. Therefore, we utilized the selectively-bred low-responder (bLR; high-anxiety) and high-responder (bHR; low-anxiety) rats to examine the effects of repeated (21-day) FGF2 treatment on CCK and FGF-R1 mRNA in the rostral VTA (VTAr). In vehicle-treated controls, both CCK and FGF-R1 mRNA levels were increased in the VTAr of bLR rats relative to bHR rats. Following FGF2 treatment, however, bHR–bLR differences in CCK and FGF-R1 mRNA expression were eliminated, due to decreased CCK mRNA levels in the VTAr of bLR rats and increased FGF-R1 expression in bHR rats. Differences after FGF2 treatment may denote distinct interactions between the CCK and FGF systems in the VTAr of bHR vs. bLR rats. Indeed, significant correlations between CCK and FGF-R1 mRNA expression were found in bHR, but not bLR rats. Colocalization studies suggest that CCK and FGF-R1 are coexpressed in some VTAr neurons. Taken together, our findings suggest that the FGF system is poised to modulate both CCK and FGF-R1 expression in the VTAr, which may be associated with individual differences in mesolimbic pathways associated with anxiety-like behavior.

Changes in expression of appetite-regulating hormones in the cunner (Tautogolabrus adspersus) during short-term fasting and winter torpor

Feeding in vertebrates is controlled by a number of appetite stimulating (orexigenic, e.g. orexin and neuropeptide Y, NPY) and appetite suppressing (anorexigenic, e.g. cholecystokinin, CCK and cocaine- and amphetamine-regulated transcript, CART) hormones. Cunners (Tautogolabrus adspersus) survive the winter in shallow coastal waters by entering a torpor-like state, during which they forgo feeding. In order to better understand the mechanisms regulating appetite/fasting in these fish, quantitative real-time PCR was used to measure transcript expression levels of four appetite-regulating hormones: NPY, CART, orexin and CCK in the forebrain (hypothalamus and telencephalon) and CCK in the gut of fed, short-term summer fasted, and natural winter torpor cunners. Summer fasting induced a decrease in hypothalamic orexin levels and telencephalon NPY, CART and CCK mRNA levels. All brain hormone mRNA levels decreased during natural torpor as compared to fed summer fish. In the gut, CCK expression levels decreased during summer fasting. These results indicate that, in cunner, orexin, NPY, CART and CCK may play a role in appetite regulation and might mediate different physiological responses to short-term summer fasting and torpor-induced long-term fasting.

Goldfish spexin: solution structure and novel function as a satiety factor in feeding control

Spexin (SPX) is a neuropeptide identified recently by bioinformatic approach. At present not much is known about its biological actions, and comparative studies of SPX in nonmammalian species are still lacking. To examine the structure and function of SPX in fish model, SPX was cloned in goldfish and found to be highly comparable with its mammalian counterparts. As revealed by NMR spectroscopies, goldfish SPX is composed of an α-helix from Gln(5) to Gln(14) with a flexible NH2 terminus from Asn(1) to Pro(4), and its molecular surface is largely hydrophobic except for Lys(11) as the only charged residue in the helical region. In goldfish, SPX transcripts were found to be widely expressed in various tissues, and protein expression of SPX was also detected in the brain. In vivo feeding studies revealed that SPX mRNA levels in the telencephalon, optic tectum, and hypothalamus of goldfish brain could be elevated by food intake. However, brain injection of goldfish SPX inhibited both basal and NPY- or orexin-induced feeding behavior and food consumption. Similar treatment also reduced transcript expression of NPY, AgRP, and apelin, with concurrent rises in CCK, CART, POMC, MCH, and CRH mRNA levels in different brain areas examined. The differential effects of SPX treatment on NPY, CCK, and MCH transcript expression could also be noted in vitro in goldfish brain cell culture. Our studies for the first time unveil the solution structure of SPX and its novel function as a satiety factor through differential modulation of central orexigenic and anorexigenic signals.

Oral administration of omega-7 palmitoleic acid induces satiety and the release of appetite-related hormones in male rats

We have analyzed the effect of palmitoleic acid on short-term food intake in male rats. Administration of omega-7 palmitoleic acid by oral gavage significantly decreased food intake compared to palmitic acid, omega-9 oleic acid, or a vehicle control. Palmitoleic acid exhibited a dose-dependent effect in this context and did not cause general malaise. A triglyceride form of palmitoleate also decreased food intake, whereas olive oil, which is rich in oleic acid, did not. Palmitoleic acid accumulated within the small intestine in a dose-dependent fashion and elevated levels of the satiety hormone cholecystokinin (CCK). Both protein and mRNA levels of CCK were affected in this context. The suppression of food intake by palmitoleic acid was attenuated by intravenous injection of devazepide, a selective peripheral CCK receptor antagonist. Palmitoleic acid did not alter the expression of peroxisome proliferator-activated receptor alpha (PPARα) target genes, and a PPARα antagonist did not affect palmitoleic acid-induced satiety. This suggests that the PPARα pathway might not be involved in suppressing food intake in response to palmitoleic acid. We have shown that orally administered palmitoleic acid induced satiety, enhanced the release of satiety hormones in rats.

The effect of fasting and refeeding on mRNA expression of PepT1 and gastrointestinal hormones regulating digestion and food intake in zebrafish (Danio rerio)

In vertebrates, a significant part of ingested protein is absorbed as di- and tripeptides through a brush border membrane proton/oligopeptide transporter protein called PepT1. The aim of the present study was to determine the effect of short-term food deprivation and refeeding in adult zebrafish (Danio rerio) on gastrointestinal mRNA expression of PepT1 as well as on the satiety hormones cholecystokinin (CCK), gastrin-releasing peptide (GRP) and ghrelin (GHR) in order to elucidate a potential mechanism driving compensatory growth. Sixty adult zebrafish were stocked in a 40-L aquarium and fed daily a commercial flake diet to satiation for 10 days where the digestive tracts (DT) of sampled fish (n = 5) were dissected out. Samplings were repeated following 1, 2 and 5 days of food deprivation and after 1, 2 and 5 days of refeeding. The RNA was extracted from all sampled DTs and analyzed by quantitative real-time PCR for the mRNA expression of PepT1, rRNA 18S, CCK, GRP and GHR. PepT1 mRNA expression increased with successive refeedings reaching a level approximately 8 times higher than pre-fast levels. CCK, GRP and GHR mRNA levels also decreased during fasting, but increased only to pre-fasting levels with refeeding. Overall, the results suggest that PepT1 may be a contributing mechanism to compensatory growth that could influence CCK secretion and GRP and GHR activity.

Response of cholecystokinin and digestive enzyme mRNA levels to various feed ingredients in yellowtail Seriola quinqueradiata

Cholecystokinin (CCK) is the key regulator hormone that stimulates the secretion of digestive pancreatic enzymes in vertebrates. In fish, little is known about the mechanism of induction of CCK in the digestive tract by feed ingredients. To investigate the response of CCK and digestive enzymes to fish feed ingredients in yellowtail Seriola quinqueradiata, we performed a number of experiments in which we measured the mRNA levels of CCK, trypsin, and lipase after oral administration of a single bolus of the ingredients. We administered fish meal and fish oil in experiment 1; high and low concentrations of fish meal in experiment 2; and five different dietary protein sources (fish meal, soybean meal, soy protein concentrate, corn gluten meal, and glutamic acid fermentation by-products) in experiment 3. In experiments 1 and 3, only fish meal significantly increased the mRNA levels of CCK and digestive enzyme. In experiment 2, a high concentration of fish meal [20 % (w/v)] significantly increased CCK and trypsin mRNA levels, but a low concentration of fish meal [1 % (w/v)] did not. These results suggest that high concentrations of fish meal (the protein source in fish feed) has the most potent effect on stimulation of CCK synthesis and secretion of digestive enzymes in yellowtail.