Recombinant Mouse Cholecystokinin receptor type A (Cckar)
Description
Table 1: Selective Ligands of CCKAR
Pancreatic β-Cell Protection
-
CCKAR activation reduces cytokine-induced β-cell apoptosis by 60–70% in mouse islets .
-
Mechanistically, β-arrestin-1 mediates anti-apoptotic effects via ERK-dependent Bcl-2 activation .
Obesity-Related Airway Hyperreactivity
-
CCKAR antagonists (e.g., devazepide) attenuate airway smooth muscle contraction in obese mice, reducing bronchoconstriction by 40% .
Central Nervous System
-
Double CCKAR/CCKBR knockout mice exhibit cortical midline defects and impaired interneuron migration .
Recombinant Production and Applications
Recombinant CCKAR is typically expressed in:
Table 2: Key Research Findings Using Recombinant CCKAR
Therapeutic Implications
-
Diabetes: CCKAR agonists enhance β-cell survival, showing promise in human islet transplants .
-
Oncology: High CCKAR expression in NSCLC correlates with brain metastasis (HR = 2.4, p < 0.01) .
-
Asthma: CCKAR inhibition reduces airway hyperreactivity in db/db mice by 35% .
Challenges and Future Directions
Product Specs
Note: We prioritize shipping the format currently in stock. However, if you have specific format requirements, please specify them in your order remarks. We will accommodate your request to the best of our ability.
Note: All our proteins are shipped with standard blue ice packs. If you require dry ice shipping, please notify us in advance as additional fees will apply.
Generally, the shelf life of liquid form is 6 months at -20°C/-80°C. The shelf life of lyophilized form is 12 months at -20°C/-80°C.
Target Background
- Chronic pancreatitis is established as an IL-33-dependent inflammation arising from synergistic interactions between the NOD1 and CCKAR signaling pathways. PMID: 26813347
- There is a functional synergy between cholecystokinin receptors CCKAR and CCKBR in mammalian brain development. PMID: 25875176
- Mice lacking CCK receptors exhibit a functional shift from the gastrin-CCK pathways to the neuronal pathway in controlling ECL cells and ultimately acid secretion. PMID: 25160855
- Cellular and subcellular localization of cholecystokinin (CCK)-1 receptors in the pancreas, gallbladder, and stomach. PMID: 25249350
- CCK-8S increases [Ca(2+)]i in gastric antral interstitial cells of Cajal via the CCK(1) receptor. PMID: 23326123
- CCK1 receptor activation, in combination with upregulated leptin signaling, does not exert a greater effect in high-fat-fed mice. PMID: 23462797
- This research suggests a role for ependymal CCK-1 receptors in infant satiety-controlling mechanisms. PMID: 23266937
- CCK-1 and -2 receptors may function synergistically in single PaPo neurons, and deletion of CCK-1 receptors could facilitate CCK-2 receptor signaling. PMID: 23038256
- This study demonstrates that cholecystokinin-1 receptor is associated with glucose homeostasis. PMID: 21871472
- Data indicate that hyperphagia in CCKR/ mice consuming a high-fat diet is reversed by blockade of the ghrelin receptor. PMID: 21277881
- Activation of neurons in the nucleus of the solitary tract following administration of T2R agonists to the GI tract involves CCK(1) and Y(2) receptors located on vagal afferent terminals in the gut wall [CCK1R]. PMID: 18003792
- The CCK-A receptor is essential for pancreatic exocrine secretion but not critical for maintaining glucose concentration and pancreatic growth in mice. PMID: 11893936
- Anxiety-related behaviors are observed in cholecystokinin-A, B, and AB receptor gene knockout mice in the plus-maze. PMID: 12459512
- Gallstone formation is enhanced in CCK-AR gene knockout mice. PMID: 12572876
- Deteriorated gallbladder contraction due to a lack of CCK-AR promotes gallstone formation after middle age. PMID: 14627338
- There are distinct roles for cholecystokinin a receptors in energy balance in rats and mice. PMID: 15123537
- Mice lacking CCK-AR exhibit larger hysteresis than mice expressing CCK-AR. PMID: 15178543
- CCKAR is involved in CCK-8S-induced depolarization of orexin neurons. PMID: 16093397
- MEKK1 likely activates a transcriptional partner of c-Jun, whose activity is maintained or enhanced in the presence of the rat cholecystokinin 1 receptor but suppressed in the presence of the mouse cholecystokinin 1 receptor. PMID: 16491099
- CCK-AR(-/-)BR(-/-) mice display lower food intake, a reduced response to exogenous ghrelin, and a lower plasma ghrelin level after fasting, although the specific receptor contributing more significantly is unknown. PMID: 17134539
- Apo A-IV and CCK(1)R play a role in PYY(3-36)-induced activation of the vagal afferent pathway and inhibition of gastric emptying, but this is likely not the pathway mediating the effects of PYY(3-36) on food intake. PMID: 17641001
- CCK-AR is an ERalpha downstream gene in the pituitary. CCK-AR may participate in the estrogen sensitization of the pituitary response to GnRH. PMID: 18000302
- These findings suggest that the CCK(1)R is involved in regulating caloric intake on a meal-to-meal basis, but other factors are responsible for regulating daily food intake. PMID: 18023701
- Data suggest a novel function of CCK-A receptors in non-image-forming photoreception, presumably through amacrine cell-mediated signal transduction pathways. PMID: 18073333
- Mice lacking CCK1Rs have fewer proliferating cells and neuroblasts compared to normal mice, and a deficiency of interneurons in the OB. PMID: 18305161
- CCK-2R is essential for responding to carbachol and producing maximal acid secretion, while the role of CCK-1R in acid secretion is less prominent. PMID: 19340558
Q&A
What are the preferred methods for detecting mouse CCKAR expression in tissue samples?
Mouse CCKAR can be detected through several complementary techniques:
-
Immunohistochemistry (IHC): Using specific antibodies (1:100 dilution, such as from Santa Cruz Biotechnology) allows visualization of CCKAR primarily in the cytoplasm and membrane. The staining can be semi-quantified using an IHC score based on staining intensity (0-3) multiplied by positive cell percentage score (1-4), with final scores ranging from 0-12 .
-
Quantitative Real-Time PCR (qRT-PCR): This method quantifies CCKAR mRNA levels using specific primers (forward: 5'-ATGGATGTGGTTGACAGCCTT-3', reverse: 5'-AAGCGTCTCATTTTCGAGCCC-3') with GAPDH as an internal control. Results are typically analyzed using the 2^-ΔΔCt method .
-
ELISA: Sandwich enzyme immunoassay kits are available with detection ranges of 0.16-10 ng/mL and sensitivity of 0.064 ng/mL. These assays use a biotin-conjugated antibody specific to mouse CCKAR followed by Avidin-HRP conjugate detection .
How does CCKAR expression differ between normal and pathological tissue conditions?
In non-small cell lung cancer (NSCLC), CCKAR expression is significantly higher in tumor tissues compared to adjacent normal tissues as demonstrated by qRT-PCR analysis . This upregulation appears to have functional significance as CCKAR has been identified as a prognostic biomarker in NSCLC and is positively associated with asynchronous brain metastasis development .
In normal physiological conditions, CCKAR shows dynamic and largely reciprocal expression patterns with CCKBR during embryonic and postnatal brain development, suggesting developmental stage-specific functions .
What is the typical sensitivity and detection range for mouse CCKAR ELISA assays?
Commercial mouse CCKAR ELISA kits typically demonstrate:
-
Sensitivity: 0.064 ng/mL
-
Detection Range: 0.16-10 ng/mL
-
Standard Concentration: 10 ng/mL
-
Assay Duration: Approximately 3.5 hours
-
Precision: Intra-assay CV% <8%; Inter-assay CV% <10%
| Concentration (ng/mL) | OD | Corrected OD |
|---|---|---|
| 10.00 | 2.018 | 1.930 |
| 5.00 | 1.794 | 1.706 |
| 2.50 | 1.102 | 1.014 |
| 1.25 | 0.837 | 0.749 |
| 0.63 | 0.539 | 0.451 |
| 0.32 | 0.312 | 0.224 |
| 0.16 | 0.216 | 0.128 |
| 0.00 | 0.088 | 0.000 |
What sample types are suitable for mouse CCKAR detection?
The following sample types have been validated for mouse CCKAR detection:
-
Tissue homogenates: Particularly from neural tissue, gastrointestinal tract, and tumors
-
Serum: With demonstrated recovery rates of 87-99%
-
EDTA plasma: With demonstrated recovery rates of >93%
-
Other biological fluids: Including cerebrospinal fluid and cell culture supernatants
Proper sample preparation is crucial, typically involving homogenization in appropriate buffers, centrifugation to remove debris, and sometimes dilution to ensure measurements fall within the assay's detection range.
How can researchers effectively analyze CCKAR G-protein coupling selectivity and promiscuity?
CCKAR serves as an ideal model for studying G protein selectivity and promiscuity due to its ability to couple with multiple G protein subtypes. Researchers can employ several approaches:
-
BRET (Bioluminescence Resonance Energy Transfer) Assays: These assays can evaluate the coupling activity of CCKAR with different G proteins (Gq, Gs, Gi, G13) by measuring energy transfer between tagged receptor and G protein components .
-
Cryo-EM Structural Analysis: Recent cryo-EM structures of sulfated CCK-8 activated CCKAR in complex with heterotrimeric Gq, Gs, or Gi proteins have revealed structural determinants responsible for G protein selectivity. All-atom root-mean-square deviation (RMSD) analyses show values of 0.84 for Gq/Gs-coupled receptors and 1.03 for Gq/Gi-coupled receptors .
-
Signaling Pathway Analysis: Researchers can differentiate coupling to:
What experimental approaches can distinguish between biased signaling pathways downstream of CCKAR?
CCKAR activates multiple signaling pathways, and researchers interested in biased signaling can:
-
Employ Pathway-Specific Biosensors: Use FRET or BRET-based biosensors to monitor real-time activation of distinct pathways (calcium, cAMP, ERK, β-arrestin recruitment).
-
Conduct Comparative Pharmacology: Test multiple ligands against pathway-specific readouts to construct signaling fingerprints and identify bias factors relative to a reference ligand (often CCK-8).
-
Utilize Phosphoproteomic Analysis: Map the complete signaling profile triggered by different CCKAR ligands to identify pathway-specific phosphorylation events.
-
Implement CRISPR-Based Pathway Interruption: Selectively disrupt individual signaling components to determine their contribution to specific cellular outcomes .
This approach is particularly relevant for developing biased CCKAR agonists that could potentially separate desired therapeutic effects (e.g., satiety) from unwanted side effects .
How can researchers effectively study the synergistic effects between CCKAR and CCKBR in neurodevelopment?
To investigate the synergistic functions of CCKAR and CCKBR in brain development:
-
Compound Knockout Models: Generate and analyze compound homozygous mutant mice lacking both CCK receptors. Previous studies have demonstrated that dual receptor loss leads to abnormalities in cortical development, including defects in midline formation, corpus callosum development, and cortical interneuron migration .
-
Comparative Transcriptome Analysis: Perform RNA-seq on embryonic neocortex from wild-type, single receptor knockouts, and double receptor knockouts to define molecular mechanisms underlying developmental defects .
-
Temporal Expression Mapping: Track the dynamic and reciprocal expression patterns of both receptors throughout embryonic and postnatal brain development using in situ hybridization and immunofluorescence .
-
Cell-Type Specific Conditional Knockouts: Generate conditional knockout models to study receptor functions in specific neural cell populations and developmental stages.
What strategies can be employed to develop and characterize positive allosteric modulators (PAMs) of CCKAR?
Researchers interested in developing PAMs for CCKAR can:
-
Conduct High-Throughput Screening: Screen chemical libraries for compounds that enhance CCK-induced responses without intrinsic agonist activity.
-
Employ Mutagenesis Studies: Identify potential allosteric binding sites through systematic mutagenesis and functional testing.
-
Perform Structural Biology Approaches: Use cryo-EM or X-ray crystallography to visualize allosteric binding pockets and ligand interactions.
-
Implement Pharmacological Characterization:
PAMs without intrinsic agonist activity represent a promising strategy for therapeutic targeting of CCKAR, as exemplified by Cinacalcet's successful application as a calcimimetic PAM for the calcium-sensing receptor .
How can researchers accurately quantify and interpret CCKAR's role in tumor progression and metastasis?
To investigate CCKAR's role in cancer:
-
Prognostic Correlation Analysis: Divide patient cohorts based on CCKAR expression levels (using determined IHC score cutoffs via ROC curve analysis) and correlate with clinical outcomes using Kaplan-Meier survival analysis and Cox-regression hazard models .
-
Metastasis Association Studies: Analyze the correlation between CCKAR expression and specific metastatic events (such as brain metastasis in NSCLC) using chi-square tests and multivariate analysis .
-
Mechanistic Investigations:
-
Perform in vitro migration/invasion assays with CCKAR overexpression or knockdown
-
Conduct in vivo metastasis models with receptor manipulation
-
Analyze downstream signaling pathways mediating metastatic potential
-
-
Therapeutic Targeting Assessment: Evaluate CCKAR-targeted agents for their ability to inhibit tumor growth and metastatic spread in preclinical models.
