CCK Receptor Research Articles

Cholecystokinin-Induced Duodenogastric Bile Reflux Increases the Severity of Indomethacin-Induced Gastric Antral Ulcers in Re-fed Mice.

We examined the involvement of cholecystokinin (CCK) in the exacerbation of indomethacin (IND)-induced gastric antral ulcers by gastroparesis caused by atropine or dopamine in mice. Male mice were fed for 2h (re-feeding) following a 22-h fast. Indomethacin (IND; 10mg/kg, s.c.) was administered after re-feeding; gastric lesions were examined 24h after IND treatment. In another experiment, mice were fed for 2h after a 22-h fast, after which the stomachs were removed 1.5h after the end of the feeding period. Antral lesions, the amount of gastric contents, and the gastric luminal bile acids concentration were measured with or without the administration of the pro- and antimotility drugs CCK-octapeptide (CCK-8), atropine, dopamine, SR57227 (5-HT3 receptor agonist), apomorphine, lorglumide (CCK1 receptor antagonist), ondansetron, and haloperidol alone and in combination. IND produced severe lesions only in the gastric antrum in re-fed mice. CCK-8, atropine, dopamine, SR57227 and apomorphine administered just after re-feeding increased bile reflux and worsened IND-induced antral lesions. These effects were significantly prevented by pretreatment with lorglumide. Although atropine and dopamine also increased the amount of gastric content, lorglumide had no effect on the delayed gastric emptying provoked by atropine and dopamine. Both ondansetron and haloperidol significantly inhibited the increase of bile reflux and the exacerbation of antral lesions induced by atropine and dopamine, respectively, but did not affect the effects of CCK-8. These results suggest that CCK-CCK1 receptor signal increases bile reflux during gastroparesis induced by atropine and dopamine, exacerbating IND-induced antral ulcers.

CKR-1 orchestrates two motor states from a single motoneuron in C. elegans

Neuromodulation is pivotal in modifying neuronal properties and motor states. CKR-1, a homolog of the cholecystokinin receptor, modulates robust escape steering and undulation body bending in C.elegans. Nevertheless, the mechanisms through which CKR-1 governs these motor states remain elusive. We elucidate the head motoneuron SMD as the orchestrator of both motor states. This regulation involves two neuropeptides: NLP-12 from DVA enhances undulation body curvature, while NLP-18 from ASI amplifies Ω-turn head curvature. Moreover, synthetic NLP-12 and NLP-18 peptides elicit CKR-1-dependent currents in Xenopus oocytes and Ca2+ transients in SMD neurons. Notably, CKR-1 shows higher sensitivity to NLP-18 compared to NLP-12. In situ patch-clamp recordings reveal CKR-1, NLP-12, and NLP-18 are not essential for neurotransmission at C.elegans neuromuscular junction, suggesting that SMD independently regulates head and body bending. Our studies illustrate that a single motoneuron SMD utilizes a cholecystokinin receptor CKR-1 to integrate two motor states.

Significant reduction of activity retention in the kidneys via optimized linker sequences in radiohybrid-based minigastrin analogs

BackgroundWe recently introduced radiohybrid (rh)-based minigastrin analogs e.g., DOTA-rhCCK-18 (DOTA-D-Dap(p-SiFA)-(D-γ-Glu)8-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2), that revealed substantially increased activity retention in the tumor. However, one major drawback of these first generation rh-based cholecystokinin-2 receptor (CCK-2R) ligands is their elevated activity levels in the kidneys, especially at later time points (24 h p.i.). Therefore, this study aimed to reduce kidney retention with regard to a therapeutic use via substitution of negatively charged D-glutamic acid moieties by hydrophilic uncharged polyethylene glycol (PEG) linkers of various length ((PEG)4 to (PEG)11). Furthermore, the influence of differently charged silicon-based fluoride acceptor (SiFA)-moieties (p-SiFA: neutral, SiFA-ipa: negatively charged, and SiFAlin: positively charged) on in vitro properties of minigastrin analogs was evaluated. Out of all compounds evaluated in vitro, the two most promising minigastrin analogs were further investigated in vivo.ResultsCCK-2R affinity of most compounds evaluated was found to be in a range of 8–20 nM (by means of apparent IC50), while ligands containing a SiFA-ipa moiety displayed elevated IC50 values. Lipophilicity was noticeably lower for compounds containing a D-γ-glutamate (D-γ-Glu) moiety next to the D-Dap(SiFA) unit as compared to their counterparts lacking the additional negative charge. Within this study, combining the most favorable CCK-2R affinity and lipophilicity, [177/natLu]Lu-DOTA-rhCCK-70 (DOTA-D-Dap(p-SiFA)-D-γ-Glu-(PEG)7-D-γ-Glu-(PEG)3-Trp-(N-Me)Nle-Asp-1-Nal-NH2; IC50: 12.6 ± 2.0 nM; logD7.4: − 1.67 ± 0.08) and [177/natLu]Lu-DOTA-rhCCK-91 (DOTA-D-Dap(SiFAlin)-D-γ-Glu-(PEG)4-D-γ-Glu-(PEG)3-Trp-(N-Me)Nle-Asp-1-Nal-NH2; IC50: 8.6 ± 0.7 nM; logD7.4 = − 1.66 ± 0.07) were further evaluated in vivo. Biodistribution data of both compounds revealed significantly reduced (p < 0.0001) activity accumulation in the kidneys compared to [177Lu]Lu-DOTA-rhCCK-18 at 24 h p.i., leading to enhanced tumor-to-kidney ratios despite lower tumor uptake. However, overall tumor-to-background ratios of the novel compounds were lower than those of [177Lu]Lu-DOTA-rhCCK-18.ConclusionWe could show that the reduction of negative charges within the linker section of radiohybrid-based minigastrin analogs led to decreased activity levels in the kidneys at 24 h p.i., while maintaining a good tumor uptake. Thus, favorable tumor-to-kidney ratios were accomplished in vivo. However, further optimization has to be done in order to improve tumor retention and general biodistribution profile.Graphical abstract

Neglected function of gastrin to reduce feeding in Siberian sturgeon (Acipenser baerii) via cholecystokinin receptor B.

Gastrin is an important intragastrointestinal hormone, but reports on its regulation of feeding behavior in fish are still scarce. This study aimed to determine the feeding regulatory function of gastrin in sturgeon. In this study, a gastrin/cholecystokinin-like peptide was identified in the genomes of sturgeon and proved to be gastrin by evolutionary tree analysis. Tissue distribution of gastrin and its receptor, cholecystokinin receptor B (CCKRB), showed that both had high mRNA abundance in the hypothalamus and gastrointestinal tract. In the duodenum, gastrin and CCKRB mRNAs were reduced at 1h of fasting, and both were also observed in the stomach and hypothalamus in response to changes in feeding status. Sulfated gastrin 17 is the major form of gastrin in vivo. Therefore, we investigated the effect of sulfated gastrin 17 on feeding by intraperitoneal injection into Siberian sturgeon using sulfated gastrin 17. The results showed that gastrin 17 significantly reduced the cumulative feeding of Siberian sturgeon in the short term (1, 3 and 6h) and long term (1, 2, 3, 4, 5 and 7days). Finally, we explored the potential mechanism of feeding inhibition after intraperitoneal injection of gastrin 17 for 7 consecutive days. The results showed that gastrin 17 treatment significantly increased the mRNA levels of anorexigenic peptides (cart, cck and pyy), while it had no significant effect on the mRNA abundance of orexigenic peptides (npy and agrp). In addition, gastrin 17 treatment significantly affected the expression of appetite signaling pathways in the hypothalamus, such that the mRNA expression of ampkα1 was significantly reduced, whereas the mRNA abundance of stat3, mtor and s6k was significantly increased. In conclusion, the present study confirmed the anorectic effect of gastrin on Siberian sturgeon.

Cholecystokinin and cholecystokinin-A receptor: An attractive treatment strategy for biliary dyskinesia?

Biliary dyskinesia is a relatively common gastrointestinal disease that is increasing in incidence as living standards improve. However, its underlying pathogenesis remains unclear, hindering the development of therapeutic drugs. Recently, "Expression and functional study of cholecystokinin-A receptors on the interstitial Cajal-like cells of the guinea pig common bile duct" demonstrated that cholecystokinin (CCK) regulates the contractile function of the common bile duct through interaction with the CCK-A receptor in interstitial Cajal-like cells, contributing to improving the academic understanding of biliary tract dynamics and providing emerging directions for the pathogenesis and clinical management of biliary dyskinesia. This letter provides a brief overview of the role of CCK and CCK-A receptors in biliary dyskinesia from the perspective of animal experiments and clinical studies, and discusses prospects and challenges for the clinical application of CCK and CCK-A receptors as potential therapeutic targets.

Molecular determinants and intracellular targets of taurine signalling in pancreatic islet β-cells.

Despite its abundance in pancreatic islets of Langerhans and proven antihyperglycemic effects, the impact of the essential amino acid, taurine, on islet β-cell biology has not yet received due consideration, which prompted the current studies exploring the molecular selectivity of taurine import into β-cells and its acute and chronic intracellular interactions. The molecular aspects of taurine transport were probed by exposing the clonal pancreatic BRIN BD11 β-cells and primary mouse and human islets to a range of the homologs of the amino acid (assayed at 2-20 mM), using the hormone release and imaging of intracellular signals as surrogate read-outs. Known secretagogues were employed to profile the interaction of taurine with acute and chronic intracellular signals. Taurine transporter TauT was expressed in the islet β-cells, with the transport of taurine and homologs having a weak sulfonate specificity but significant sensitivity to the molecular weight of the transporter. Taurine, hypotaurine, homotaurine, and β-alanine enhanced insulin secretion in a glucose-dependent manner, an action potentiated by cytosolic Ca2+ and cAMP. Acute and chronic β-cell insulinotropic effects of taurine were highly sensitive to co-agonism with GLP-1, forskolin, tolbutamide, and membrane depolarization, with an unanticipated indifference to the activation of PKC and CCK8 receptors. Pre-culturing with GLP-1 or KATP channel inhibitors sensitized or, respectively, desensitized β-cells to the acute taurine stimulus. Together, these data demonstrate the pathways whereby taurine exhibits a range of beneficial effects on insulin secretion and β-cell function, consistent with the antidiabetic potential of its dietary low-dose supplementation.

Precision peptide theranostics: developing N- to C-terminus optimized theranostics targeting cholecystokinin-2 receptor.

Peptides are ideal for theranostic development as they afford rapid target accumulation, fast clearance from background tissue, and exhibit good tissue penetration. Previously, we developed a novel series of peptides that presented discreet folding propensity leading to an optimal candidate [68Ga]Ga-DOTA-GA1 ([D-Glu]6-Ala-Tyr-NMeGly-Trp-NMeNle-Asp-Nal-NH2) with 50 pM binding affinity against cholecystokinin-2 receptors (CCK2R). However, we were confronted with challenges of unfavorably high renal uptake. Methods: A structure activity relationship study was undertaken of the lead theranostic candidate. Prudent structural modifications were made to the peptide scaffold to evaluate the contributions of specific N-terminal residues to the overall biological activity. Optimal candidates were then evaluated in nude mice bearing transfected A431-CCK2 tumors, and their biodistribution was quantitated ex vivo. Results: We identified and confirmed that D-Glu3 to D-Ala3 substitution produced 2 optimal candidates, [68Ga]Ga-DOTA-GA12 and [68Ga]Ga-DOTA-GA13. These radiopeptides presented with high target/background ratios, enhanced tumor retention, excellent metabolic stability in plasma and mice organ homogenates, and a 4-fold reduction in renal uptake, significantly outperforming their non-alanine counterparts. Conclusions: Our study identified novel radiopharmaceutical candidates that target the CCK2R. Their high tumor uptake and reduced renal accumulation warrant clinical translation.

Inter-Organ Communication Involved in Brown Adipose Tissue Thermogenesis.

Brown and beige adipocytes produce heat from substrates such as fatty acids and glucose. Such heat productions occur in response to various stimuli and are called adaptive non-shivering thermogenesis. This review introduces mechanisms known to regulate brown and beige adipocyte thermogenesis. Leptin and fibroblast growth factor 21 (FGF21) are examples of periphery-derived humoral factors that act on the central nervous system (CNS) and increase brown adipose tissue (BAT) thermogenesis. Additionally, neuronal signals such as those induced by intestinal cholecystokinin and hepatic peroxisome proliferator-activated receptor γ travel through vagal afferent-CNS-sympathetic efferent-BAT pathways and increase BAT thermogenesis. By contrast, some periphery-derived humoral factors (ghrelin, adiponectin, plasminogen activator inhibitor-1, and soluble leptin receptor) act also on CNS but inhibit BAT thermogenesis. Neuronal signals also reduce BAT sympathetic activities and BAT thermogenesis, one such example being signals derived by hepatic glucokinase activation. Beige adipocytes can be induced by myokines (interleukin 6, irisin, and β-aminoisobutyric acid), hepatokines (FGF21), and cardiac-secreted factors (brain natriuretic peptide). Cold temperature and leptin also stimulate beige adipocytes via sympathetic activation. Further investigation on inter-organ communication involving adipocyte thermogenesis may lead to the elucidation of how body temperature is regulated and, moreover, to the development of novel strategies to treat metabolic disorders.

Cholecystokinin-A signaling regulates automaticity of pacemaker cardiomyocytes.

Aims: The behavior of pacemaker cardiomyocytes (PCs) in the sinoatrial node (SAN) is modulated by neurohormonal and paracrine factors, many of which signal through G-protein coupled receptors (GPCRs). The aims of the present study are to catalog GPCRs that are differentially expressed in the mammalian SAN and to define the acute physiological consequences of activating the cholecystokinin-A signaling system in isolated PCs. Methods and results: Using bulk and single cell RNA sequencing datasets, we identify a set of GPCRs that are differentially expressed between SAN and right atrial tissue, including several whose roles in PCs and in the SAN have not been thoroughly characterized. Focusing on one such GPCR, Cholecystokinin-A receptor (CCKAR), we demonstrate expression of Cckar mRNA specifically in mouse PCs, and further demonstrate that subsets of SAN fibroblasts and neurons within the cardiac intrinsic nervous system express cholecystokinin, the ligand for CCKAR. Using mouse models, we find that while baseline SAN function is not dramatically affected by loss of CCKAR, the firing rate of individual PCs is slowed by exposure to sulfated cholecystokinin-8 (sCCK-8), the high affinity ligand for CCKAR. The effect of sCCK-8 on firing rate is mediated by reduction in the rate of spontaneous phase 4 depolarization of PCs and is mitigated by activation of beta-adrenergic signaling. Conclusion: (1) PCs express many GPCRs whose specific roles in SAN function have not been characterized, (2) Activation of the cholecystokinin-A signaling pathway regulates PC automaticity.

TRPV1 enhances cholecystokinin signaling in primary vagal afferent neurons and mediates the central effects on spontaneous glutamate release in the NTS.

The gut peptide cholecystokinin (CCK) is released during feeding and promotes satiation by increasing excitation of vagal afferent neurons that innervate the upper gastrointestinal tract. Vagal afferent neurons express CCK1 receptors (CCK1Rs) in the periphery and at central terminals in the nucleus of the solitary tract (NTS). While the effects of CCK have been studied for decades, CCK receptor signaling and coupling to membrane ion channels are not entirely understood. Previous findings have implicated L-type voltage-gated calcium channels as well as transient receptor potential (TRP) channels in mediating the effects of CCK, but the lack of selective pharmacology has made determining the contributions of these putative mediators difficult. The nonselective ion channel transient receptor potential vanilloid subtype 1 (TRPV1) is expressed throughout vagal afferent neurons and controls many forms of signaling, including spontaneous glutamate release onto NTS neurons. Here we tested the hypothesis that CCK1Rs couple directly to TRPV1 to mediate vagal signaling using fluorescent calcium imaging and brainstem electrophysiology. We found that CCK signaling at high concentrations (low-affinity binding) was potentiated in TRPV1-containing afferents and that TRPV1 itself mediated the enhanced CCK1R signaling. While competitive antagonism of TRPV1 failed to alter CCK1R signaling, TRPV1 pore blockade or genetic deletion (TRPV1 KO) significantly reduced the CCK response in cultured vagal afferents and eliminated its ability to increase spontaneous glutamate release in the NTS. Together, these results establish that TRPV1 mediates the low-affinity effects of CCK on vagal afferent activation and control of synaptic transmission in the brainstem.NEW & NOTEWORTHY Cholecystokinin (CCK) signaling via the vagus nerve reduces food intake and produces satiation, yet the signaling cascades mediating these effects remain unknown. Here we report that the capsaicin receptor transient receptor potential vanilloid subtype 1 (TRPV1) potentiates CCK signaling in the vagus and mediates the ability of CCK to control excitatory synaptic transmission in the nucleus of the solitary tract. These results may prove useful in the future development of CCK/TRPV1-based therapeutic interventions.

Elevated Serum Gastrin Is Associated with Melanoma Progression: Putative Role in Increased Migration and Invasion of Melanoma Cells

Micro-environmental factors, including stromal and immune cells, cytokines, and circulating hormones are well recognized to determine cancer progression. Melanoma cell growth was recently shown to be suppressed by cholecystokinin/gastrin (CCK) receptor antagonists, and our preliminary data suggested that melanoma patients with Helicobacter gastritis (which is associated with elevated serum gastrin) might have an increased risk of cancer progression. Therefore, in the present study, we examined how gastrin may act on melanoma cells. In 89 melanoma patients, we found a statistically significant association between circulating gastrin concentrations and melanoma thickness and metastasis, which are known risk factors of melanoma progression and prognosis. Immunocytochemistry using a validated antibody confirmed weak to moderate CCK2R expression in both primary malignant melanoma cells and the melanoma cell lines SK-MEL-2 and G361. Furthermore, among the 219 tumors in the Skin Cutaneous Melanoma TCGA Pan-Cancer dataset showing gastrin receptor (CCKBR) expression, significantly higher CCKBR mRNA levels were linked to stage III–IV than stage I–II melanomas. In both cell lines, gastrin increased intracellular calcium levels and stimulated cell migration and invasion through mechanisms inhibited by a CCK2 receptor antagonist. Proteomic studies identified increased MMP-2 and reduced TIMP-3 levels in response to gastrin that were likely to contribute to the increased migration of both cell lines. However, the effects of gastrin on tumor cell invasion were relatively weak in the presence of the extracellular matrix. Nevertheless, dermal fibroblasts/myofibroblasts, known also to express CCK2R, increased gastrin-induced cancer cell invasion. Our data suggest that in a subset of melanoma patients, an elevated serum gastrin concentration is a risk factor for melanoma tumor progression, and that gastrin may act on both melanoma and adjacent stromal cells through CCK2 receptors to promote mechanisms of tumor migration and invasion.

Intestinal microbiome dysbiosis in alcohol-dependent patients and its effect on rat behaviors.

Intestinal microbiome dysbiosis is associated with psychiatric disease through the "microbiota-gut-brain" axis. Here, we revealed that there was obvious intestinal microbiome (including bacterial and fungal) dysbiosis in alcohol-dependent patients. Alcohol consumption seriously disturbs the gut equilibrium between bacteria and fungi, reduces the interactions among bacterial-fungal trans-kingdom, and increases intestinal permeability. Gut microbiota should be considered as a whole to study the development of alcohol dependence. The gut microbiome of alcohol-dependent patients increased the anxiety- and depression-like behavior in rats. The gut microbiota dysbiosis may promote the development of alcohol dependence by regulating the endogenous cholecystokinin (CCK) and related receptors. Hence, regulating the balance of gut microbiota and the endogenous CCK may be a potential strategy for reducing the risk of relapse in alcohol addiction patients.

Substance P, NPY, CCK and their receptors in five brain regions in major depressive disorder with transcriptomic analysis of locus coeruleus neurons

Major depressive disorder (MDD) is a serious disease and a burden to patients, families and society. Rodent experiments and human studies suggest that several neuropeptide systems are involved in mood regulation. The aim of this study is two-fold: (i) to monitor, with qPCR, transcript levels of the substance P/tachykinin (TAC), NPY and CCK systems in bulk samples from control and suicide subjects, targeting five postmortem brain regions including locus coeruleus (LC); and (ii) to analyse expression of neuropeptide family transcripts in LC neurons of ‘normal’ postmortem brains by using laser capture microdissection with Smart-Seq2 RNA sequencing. qPCR revealed distinct regional expression patterns in male and female controls with higher levels for the TAC system in the dorsal raphe nucleus and LC, versus higher transcripts levels of the NPY and CCK systems in prefrontal cortex. In suicide patients, TAC, TAC receptors and a few NPY family transcript levels were increased mainly in prefrontal cortex and LC. The second study on ‘normal’ noradrenergic LC neurons revealed expression of transcripts for GAL, NPY, TAC1, CCK, and TACR1 and many other peptides (e.g. Cerebellin4 and CARTPT) and receptors (e.g. Adcyap1R1 and GPR173). These data and our previous results on suicide brains indicates that the tachykinin and galanin systems may be valid targets for developing antidepressant medicines. Moreover, the perturbation of neuropeptide systems in MDD patients, and the detection of further neuropeptide and receptor transcripts in LC, shed new light on signalling in noradrenergic LC neurons and on mechanisms possibly associated with mood disorders.

Expression and functional study of cholecystokinin-A receptors on the interstitial Cajal-like cells of the guinea pig common bile duct.

Many studies have shown that interstitial Cajal-like cell (ICLC) abnormalities are closely related to a variety of dynamic gastrointestinal disorders. ICLCs are pacemaker cells for gastrointestinal movement and are involved in the transmission of nerve impulses. To elucidate the expression profile and significance of cholecystokinin-A (CCK-A) receptors in ICLCs in the common bile duct (CBD), as well as the role of CCK in regulating CBD motility through CCK-A receptors on CBD ICLCs. The levels of tyrosine kinase receptor (c-kit) and CCK-A receptors in CBD tissues and isolated CBD cells were quantified using the double immunofluorescence labeling technique. The CCK-mediated enhancement of the movement of CBD muscle strips through CBD ICLCs was observed by a muscle strip contraction test. Immunofluorescence showed co-expression of c-kit and CCK-A receptors in the CBD muscularis layer. Observations of isolated CBD cells showed that c-kit was expressed on the surface of ICLCs, the cell body and synapse were colored and polygonal, and some cells presented protrusions and formed networks adjacent to the CBD while others formed filaments at the synaptic terminals of local cells. CCK-A receptors were also expressed on CBD ICLCs. At concentrations ranging from 10-6 mol/L to 10-10 mol/L, CCK promoted CBD smooth muscle contractility in a dose-dependent manner. In contrast, after ICLC removal, the contractility mediated by CCK in CBD smooth muscle decreased. CCK-A receptors are highly expressed on CBD ICLCs, and CCK may regulate CBD motility through the CCK-A receptors on ICLCs.

CCK-8 enhances acid-sensing ion channel currents in rat primary sensory neurons

Cholecystokinin (CCK) is a peptide that has been implicated in pain modulation. Acid sensitive ion channels (ASICs) also play an important role in pain associated with tissue acidification. However, it is still unclear whether there is an interaction between CCK signaling and ASICs during pain process. Herein, we report that a functional link between them in rat dorsal root ganglion (DRG) neurons. Pretreatment with CCK-8 concentration-dependently increased acid-evoked ASIC currents. CCK-8 increased the maximum response of ASICs to acid, but did not changed their acid sensitivity. Enhancement of ASIC currents by CCK-8 was mediated by the stimulation of CCK2 receptor (CCK2R), rather than CCK1R. The enhancement of ASIC currents by CCK-8 was prevented by application of either G-protein inhibitor GDP-β-S or protein kinase C (PKC) inhibitor GF109203×, but not by protein kinase A (PKA) inhibitor H-89 or JNK inhibitor SP600125. Moreover, CCK-8 increased the number of action potentials triggered by acid stimuli by activating CCK2R. Finally, CCK-8 dose-dependently exacerbated acid-induced nociceptive behavior in rats through local CCK2R. Together, these results indicated that CCK-8/CCK2R activation enhanced ASIC-mediated electrophysiological activity in DRG neurons and nociception in rats. The enhancement effect depended on G-proteins and intracellular PKC signaling rather than PKA and JNK signaling pathway. These findings provided that CCK-8/CCK2R is an important therapeutic target for ASIC-mediated pain.

CCK2-receptor deficiency impairs immune balance by influencing CD4+ T cells development by inhibiting cortical-thymic-epithelial-cells.

Immune balance is crucial for an organism's survival and is inseparable from the regulation of the nervous system. Accumulating evidence indicates that cholecystokinin (CCK) plays an important role in mediating the immune response through the activation of cholecystokinin receptors (CCKRs). However, it remains unclear whether CCKRs deficiency may impair immune balance. Here, we showed that CCK2R-deficient adult mice were immunocompromised and had an increased risk of shock and even death in an endotoxemia (ETM)/endotoxin shock (ES) model. In addition, in both adult and juvenile mice, CCK2R deficiency not only influenced the development of CD4 single-positive (SP) thymocytes in thymic positive selection but also decreased the population of CD3+ CD4+ T cells in the spleen. More importantly, CCK2R deficiency inhibited the expression of major histocompatibility complex class II (MHC II) and CD83 on cortical thymic epithelial cells (cTECs) in juvenile and adult mice. Overall, our study suggests that CCK2R is essential for maintaining CD4+ T cell development in the thymus and reveals that CCK2R plays an important role in maintaining immune balance.

Effect of the dipeptide retro-analog of cholecystokinin tetrapeptide (GB-115) on the behavior of rhesus monkeys in isolation

Introduction. The developed domestic dipeptide retro-analog of cholecystokinin (CCK) tetrapeptide (N-(6-phenylhexanoyl)-glycyltryptophan amide, compound GB-115) with antagonistic properties towards CCK1 receptors has anxiolytic activity, previously shown in preclinical and clinical studies. Purpose of the study. Evaluation of the anxiolytic effect of GB-115 in tablet dosage form with subchronic oral administration in comparison with phenazepam in laboratory primates. Methods. The experiment was performed on four male rhesus monkeys (Macaca mulatta) aged 5.7–6.7 years. After a 30-day period of adaptation to the conditions of individual housing, an experiment was performed with GB-115 (0.001 g tablets) and then with phenazepam (0.0005 g tablets). Both drugs were given one at a time (7 days), and then 2 tablets (7 days). Behavior was assessed by observing the object with registration according to the “Yes-No” principle of ethogram elements in the background periods, during the administration of drugs and during their withdrawal. Using enzyme immunoassay, the content of stress response biomarkers: cortisol and dehydroepiandrosterone sulfate (DHEA-S) was determined in blood serum. Results. GB-115 (0.001 g each) and phenazepam (0.001 g each) reduced the stay of animals in the upper part of the cage compared to the background period, which indicates a decrease in the stress response. GB-115 (0.002 g each) decreased the cortisol/DHEA-S ratio. Phenazepam dose-dependently reduced serum cortisol levels without affecting DHEA-S levels; with the administration of phenazepam (0.001 g), a decrease in the cortisol/DHEA-S ratio was also recorded. Conclusion. A positive effect of GB-115, when administered subchronically, on the weakening of the emotional stress reaction and restoration of adaptive behavior in rhesus monkeys was revealed, comparable to the effect of phenazepam, which confirms the possibility of using blockade of CCK1 receptors as one of the approaches for the treatment of anxiety disorders.

Gut-derived peptide hormone receptor expression in the developing mouse hypothalamus.

In adult organisms, a number of receptors have been identified which modulate metabolic processes related to peptides derived from the intestinal tract. These receptors play significant roles in glucose homeostasis, food intake and energy balance. Here we assess these classical metabolic receptors and their expression as well as their potential role in early development of hypothalamic neuronal circuits. Chow-fed C57BL6/N female mice were mated and hypothalamic tissue was collected from offspring across postnatal development (postnatal day 7-21). Subsequent qPCR and Western Blot analyses were used to determine mRNA and protein changes in gut-derived peptide hormone receptors. Correlations to body weight, blood glucose and circulating leptin levels were analyzed. We describe the gene expression and dynamic protein regulation of key gut-derived peptide hormone receptors in the early postnatal period of the mouse brain. Specifically, we show changes to Gastric inhibitory polypeptide receptor (GIPR), glucagon-like peptide 1 receptor (GLP1R), and cholecystokinin receptor 2 (CCK2R) in the developing hypothalamus. The changes to GIPR and InsR seem to be strongly negatively correlated with body weight. This comprehensive analysis underscores the need to understand the roles of maternal-derived circulating gut hormones and their direct effect on offspring brain development.

Effect of cholecystokinin on small intestinal motility in suncus murinus

In a fasting gastrointestinal tract, a characteristic cyclical rhythmic migrating motor complex (MMC) occur that comprises of three phases: I, II, and III. Among these, phase III contractions propagate from the stomach to the lower intestine in mammals, including humans, dogs, and Suncus murinus (suncus). Apart from the phase III of MMC propagating from the stomach, during the gastric phase II, small intestine-originated strong contractions propagate to the lower small intestine; however, the mechanism of contractions originating in the small intestine has not been clarified. In this study, we aimed to elucidate the role of cholecystokinin (CCK) in small intestinal motility. Administration of sulfated CCK-8 in phase I induced phase II-like contractions in the small intestine, which lasted for approximately 10–20 min and then returned to the baseline, while no change was observed in the stomach. Contractions of small intestine induced by CCK-8 were abolished by lorglumide, a CCK1 receptor antagonist. Gastrin, a ligand for the CCK2 receptor, evoked strong contractions in the stomach, but did not induce contractions in the small intestine. To examine the effect of endogenous CCK on contractions of small intestinal origin, lorglumide was administered during phase II. However, there was no change in the duodenal motility pattern, and strong contractions of small intestinal origin were not abolished by treatment with lorglumide. These results suggest that exogenous CCK stimulates contractions of small intestine via CCK1 receptors, whereas endogenous CCK is not involved in the strong contractions of small intestinal origin.

Development of Highly Potent Clinical Candidates for Theranostic Applications against Cholecystokinin-2 Receptor Positive Cancers.

Peptide receptor radionuclide therapy (PRRT) is a promising form of systemic radiation therapy designed to eradicate cancer. Cholecystokinin-2 receptor (CCK2R) is an important molecular target that is highly expressed in a range of cancers. This study describes the synthesis and in vivo characterization of a novel series of 177Lu-labeled peptides ([177Lu]Lu-2b-4b) in comparison with the reference CCK2R-targeting peptide CP04 ([177Lu]Lu-1b). [177Lu]Lu-1b-4b showed high chemical purity (HPLC ≥ 94%), low Log D7.4 (-4.09 to -4.55) with strong binding affinity to CCK2R (KD 0.097-1.61 nM), and relatively high protein binding (55.6-80.2%) and internalization (40-67%). Biodistribution studies of the novel 177Lu-labeled peptides in tumors (AR42J and A431-CCK2R) showed uptake one- to eight-fold greater than the reference compound CP04 at 1, 24, and 48 h. Rapid clearance and high tumor uptake and retention were established for [177Lu]Lu-2b-4b, making these compounds excellent candidates for theranostic applications against CCK2R-expressing tumors.