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Neuropharmacology Jun 2020Gastrin-releasing peptide (GRP) receptor-expressing (GRPR) neurons have a central role in the spinal transmission of itch. Because their fundamental regulatory...
Gastrin-releasing peptide (GRP) receptor-expressing (GRPR) neurons have a central role in the spinal transmission of itch. Because their fundamental regulatory mechanisms are not yet understood, it is important to determine how such neurons are excited and integrate itch sensation. In this study, we investigated the mechanisms for the activation of itch-responsive GRPR neurons in the spinal dorsal horn (SDH). GRPR neurons expressed the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) containing the GluR2 subunit. In mice, peripherally elicited histaminergic and non-histaminergic itch was prevented by intrathecal (i.t.) administration of the AMPAR antagonist NBQX, which was consistent with the fact that firing of GRPR neurons in SDH under histaminergic and non-histaminergic itch was completely blocked by NBQX, but not by the GRPR antagonist RC-3095. Because GRP neurons in SDH contain glutamate, we investigated the role of GRP (GRP/Glu) neurons in regulating itch. Chemogenetic inhibition of GRP neurons suppressed both histaminergic and non-histaminergic itch without affecting the mechanical pain threshold. In nonhuman primates, i.t. administration of NBQX also attenuated peripherally elicited itch without affecting the thermal pain threshold. In a mouse model of diphenylcyclopropenone (DCP)-induced contact dermatitis, GRP, GRPR, and AMPAR subunits were upregulated in SDH. DCP-induced itch was prevented by either silencing GRP neurons or ablation of GRPR neurons. Altogether, these findings demonstrate that GRP and glutamate cooperatively regulate GRPR AMPAR neurons in SDH, mediating itch sensation. GRP-GRPR and the glutamate-AMPAR system may play pivotal roles in the spinal transmission of itch in rodents and nonhuman primates.
Topics: Animals; Bombesin; Cyclopropanes; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Macaca mulatta; Male; Mice; Mice, Inbred C57BL; Mice, Inbred ICR; Neurons; Peptide Fragments; Pruritus; Receptors, AMPA; Receptors, Bombesin; Spinal Cord Dorsal Horn
PubMed: 32142790
DOI: 10.1016/j.neuropharm.2020.108025 -
The Journal of Comparative Neurology Nov 2022Gastrin-releasing peptide (GRP) and its receptor (GRPR) have been identified as itch mediators in the spinal and trigeminal somatosensory systems in rodents. In...
Characterization of the expression of gastrin-releasing peptide and its receptor in the trigeminal and spinal somatosensory systems of Japanese macaque monkeys: Insight into humans.
Gastrin-releasing peptide (GRP) and its receptor (GRPR) have been identified as itch mediators in the spinal and trigeminal somatosensory systems in rodents. In primates, there are few reports of GRP/GRPR expression or function in the spinal sensory system and virtually nothing is known in the trigeminal system. The aim of the present study was to characterize GRP and GRPR in the trigeminal and spinal somatosensory system of Japanese macaque monkeys (Macaca fuscata). cDNA encoding GRP was isolated from the macaque dorsal root ganglion (DRG) and exhibited an amino acid sequence that was highly conserved among mammals and especially in primates. Immunohistochemical analysis demonstrated that GRP was expressed mainly in the small-sized trigeminal ganglion and DRG in adult macaque monkeys. Densely stained GRP-immunoreactive (ir) fibers were observed in superficial layers of the spinal trigeminal nucleus caudalis (Sp5C) and the spinal cord. In contrast, GRP-ir fibers were rarely observed in the principal sensory trigeminal nucleus and oral and interpolar divisions of the spinal trigeminal nucleus. cDNA cloning, in situ hybridization, and Western blot revealed substantial expression of GRPR mRNA and GRPR protein in the macaque spinal dorsal horn and Sp5C. Our Western ligand blot and ligand derivative stain for GRPR revealed that GRP directly bound in the macaque Sp5C and spinal dorsal horn as reported in rodents. Finally, GRP-ir fibers were also detected in the human spinal dorsal horn. The spinal and trigeminal itch neural circuits labeled with GRP and GRPR appear to function also in primates.
Topics: Animals; DNA, Complementary; Gastrin-Releasing Peptide; Humans; Ligands; Macaca fuscata; Pruritus; Receptors, Bombesin; Sense Organs
PubMed: 35686563
DOI: 10.1002/cne.25376 -
Proceedings of the National Academy of... Feb 2023Gastrin releasing peptide receptor (GRPR), a member of the bombesin (BBN) G protein-coupled receptors, is aberrantly overexpressed in several malignant tumors, including...
Gastrin releasing peptide receptor (GRPR), a member of the bombesin (BBN) G protein-coupled receptors, is aberrantly overexpressed in several malignant tumors, including those of the breast, prostate, pancreas, lung, and central nervous system. Additionally, it also mediates non-histaminergic itch and pathological itch conditions in mice. Thus, GRPR could be an attractive target for cancer and itch therapy. Here, we report the inactive state crystal structure of human GRPR in complex with the non-peptide antagonist PD176252, as well as two active state cryo-electron microscopy (cryo-EM) structures of GRPR bound to the endogenous peptide agonist gastrin-releasing peptide and the synthetic BBN analog [D-Phe, β-Ala, Phe, Nle] Bn (6-14), in complex with G heterotrimers. These structures revealed the molecular mechanisms for the ligand binding, receptor activation, and G proteins signaling of GRPR, which are expected to accelerate the structure-based design of GRPR antagonists and agonists for the treatments of cancer and pruritus.
Topics: Male; Humans; Mice; Animals; Receptors, Bombesin; Cryoelectron Microscopy; Bombesin; Gastrin-Releasing Peptide; Neoplasms; Pruritus
PubMed: 36724251
DOI: 10.1073/pnas.2216230120 -
Journal of Diabetes Research 2022Abdominal obesity coupled with polygenic hereditary defects is considered the initial event in the development of metabolic syndrome (MS). The purpose of this study was...
Abdominal obesity coupled with polygenic hereditary defects is considered the initial event in the development of metabolic syndrome (MS). The purpose of this study was to analyse the frequency with which polymorphic loci of adiponectin () and leptin () genes occur in patients with MS and the association between the symptoms of MS and these polymorphisms. DNA was isolated from the whole blood of 207 patients with MS and 100 healthy individuals (control group) using the phenol-chloroform method. Gene polymorphisms were determined using real-time polymerase chain reaction (PCR). The most common variant of the (rs2241766) gene among MS patients was the GT genotype. The A allele of the (rs7799039) gene was found to be the most frequent in MS patients. The highest systolic blood pressure was found in carriers of the GG genotype of the (rs7799039) gene. The carriers of the (rs2241766) GT genotype were associated with the highest systolic blood pressure and (); carriers of the (rs2241766) GG genotype were associated with the highest diastolic blood pressure, hyperglycaemia, and elevated glycated haemoglobin (HbA1c). The results of this study allowed us to establish the unique gene variants associated with the risk of developing MS in the Crimean population.
Topics: Adiponectin; Chloroform; Glycated Hemoglobin; Humans; Leptin; Metabolic Syndrome; Phenols; Polymorphism, Single Nucleotide; Receptors, Bombesin; Receptors, Colony-Stimulating Factor; Receptors, Formyl Peptide; Receptors, Leptin; Ryanodine Receptor Calcium Release Channel
PubMed: 36117520
DOI: 10.1155/2022/9881422 -
British Journal of Cancer Jan 2000Bombesin-like peptides and their receptors are widely distributed throughout the gut and are potential mitogens for a number of gastrointestinal (GI) cancers. We have...
Bombesin-like peptides and their receptors are widely distributed throughout the gut and are potential mitogens for a number of gastrointestinal (GI) cancers. We have analysed the expression of bombesin-like peptides and their receptor subtypes in normal and neoplastic colorectal tissue. Expression was analysed by reverse transcription polymerase chain reaction (RT-PCR) using receptor and ligand subtype-specific primers and then expression localized by in situ hybridization (ISH) with riboprobes synthesized by in vitro transcription of cloned PCR product. Colorectal cancer tissue and matched normal mucosa from 23 patients were studied. Two of these patients had synchronous adenomatous polyps and two had synchronous hepatic metastases which were also studied. An additional two patients with adenomatous polyps were studied along with matched normal mucosa. Gastrin releasing peptide (GRP) receptor and ligand expression was present in all samples but with overall greater expression in the tumour samples. Neuromedin B (NMB) receptor expression was not detectable. NMB ligand was detected in all but one mucosal sample with overall overexpression in the tumour samples. Bombesin receptor subtype 3 (BRS-3) receptor expression was not detectable. These data support the possibility that GRP may be an autocrine growth factor in colorectal cancer.
Topics: Adenocarcinoma; Colon; Colonic Neoplasms; Gene Expression; Humans; In Situ Hybridization; Intestinal Mucosa; Ligands; Neoplasm Proteins; Neurokinin B; RNA, Messenger; Receptors, Bombesin; Rectal Neoplasms; Rectum; Reverse Transcriptase Polymerase Chain Reaction
PubMed: 10638978
DOI: 10.1054/bjoc.1998.0888 -
PloS One 2017Physical measurement of tumor volume reduction is the most commonly used approach to assess tumor progression and treatment efficacy in mouse tumor models. However, it... (Comparative Study)
Comparative Study
Physical measurement of tumor volume reduction is the most commonly used approach to assess tumor progression and treatment efficacy in mouse tumor models. However, it is relatively insensitive, and often requires long treatment courses to achieve gross physical tumor destruction. As alternatives, several non-invasive imaging methods such as bioluminescence imaging (BLI), fluorescence imaging (FLI) and positron emission tomography (PET) have been developed for more accurate measurement. As tumors have elevated glucose metabolism, 18F-fludeoxyglucose (18F-FDG) has become a sensitive PET imaging tracer for cancer detection, diagnosis, and efficacy assessment by measuring alterations in glucose metabolism. In particular, the ability of 18F-FDG imaging to detect drug-induced effects on tumor metabolism at a very early phase has dramatically improved the speed of decision-making regarding treatment efficacy. Here we demonstrated an approach with FLI that offers not only comparable performance to PET imaging, but also provides additional benefits, including ease of use, imaging throughput, probe stability, and the potential for multiplex imaging. In this report, we used sorafenib, a tyrosine kinase inhibitor clinically approved for cancer therapy, for treatment of a mouse tumor xenograft model. The drug is known to block several key signaling pathways involved in tumor metabolism. We first identified an appropriate sorafenib dose, 40 mg/kg (daily on days 0-4 and 7-10), that retained ultimate therapeutic efficacy yet provided a 2-3 day window post-treatment for imaging early, subtle metabolic changes prior to gross tumor regression. We then used 18F-FDG PET as the gold standard for assessing the effects of sorafenib treatment on tumor metabolism and compared this to results obtained by measurement of tumor size, tumor BLI, and tumor FLI changes. PET imaging showed ~55-60% inhibition of tumor uptake of 18F-FDG as early as days 2 and 3 post-treatment, without noticeable changes in tumor size. For comparison, two FLI probes, BombesinRSense™ 680 (BRS-680) and Transferrin-Vivo™ 750 (TfV-750), were assessed for their potential in metabolic imaging. Metabolically active cancer cells are known to have elevated bombesin and transferrin receptor levels on the surface. In excellent agreement with PET imaging, the BRS-680 imaging showed 40% and 79% inhibition on days 2 and 3, respectively, and the TfV-750 imaging showed 65% inhibition on day 3. In both cases, no significant reduction in tumor volume or BLI signal was observed during the first 3 days of treatment. These results suggest that metabolic FLI has potential preclinical application as an additional method for detecting drug-induced metabolic changes in tumors.
Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Colonic Neoplasms; Disease Progression; Dose-Response Relationship, Drug; Fluorescent Dyes; Fluorodeoxyglucose F18; Humans; Mice, Transgenic; Molecular Imaging; Neoplasm Transplantation; Niacinamide; Optical Imaging; Phenylurea Compounds; Positron-Emission Tomography; Radiopharmaceuticals; Random Allocation; Receptors, Bombesin; Receptors, Transferrin; Sorafenib; Treatment Outcome; Tumor Burden
PubMed: 28792505
DOI: 10.1371/journal.pone.0182689 -
Proceedings. Biological Sciences Oct 2022The neural bases of itchy eye transmission remain unclear compared with those involved in body itch. Here, we show in rodents that the gastrin-releasing peptide receptor...
The neural bases of itchy eye transmission remain unclear compared with those involved in body itch. Here, we show in rodents that the gastrin-releasing peptide receptor (GRPR) of the trigeminal sensory system is involved in the transmission of itchy eyes. Interestingly, we further demonstrate a difference in scratching behaviour between the left and right hindfeet in rodents; histamine instillation into the conjunctival sac of both eyes revealed right-foot biased laterality in the scratching movements. Unilateral histamine instillation specifically induced neural activation in the ipsilateral sensory pathway, with no significant difference between the activations following left- and right-eye instillations. Thus, the behavioural laterality is presumably due to right-foot preference in rodents. Genetically modified rats with specific depletion of expressing neurons in the trigeminal sensory nucleus caudalis of the medulla oblongata exhibited fewer and shorter histamine-induced scratching movements than controls and eliminated the footedness. These results taken together indicate that the -expressing neurons are required for the transmission of itch sensation from the eyes, but that foot preference is generated centrally. These findings could open up a new field of research on the mechanisms of the laterality in vertebrates and also offer new potential therapeutic approaches to refractory pruritic eye disorders.
Topics: Animals; Rats; Histamine; Pruritus; Receptors, Bombesin; Functional Laterality; Eye
PubMed: 36259204
DOI: 10.1098/rspb.2022.1126 -
The Journal of Biological Chemistry Jul 2001Substance P analogues including [d-Arg1,d-Phe5,d-Trp7,9,Leu11]substance P (SpD) act as "broad spectrum neuropeptide antagonists" and are potential anticancer agents that...
Substance P analogues including [d-Arg1,d-Phe5,d-Trp7,9,Leu11]substance P (SpD) act as "broad spectrum neuropeptide antagonists" and are potential anticancer agents that inhibit the growth of small cell lung cancer cells in vitro and in vivo. However, their mechanism of action is controversial and not fully understood. Although these compounds block bombesin-induced mitogenesis and signal transduction, they also have agonist activity. The mechanism underlying this agonist activity was examined. SpD binds to the ligand-binding site of the bombesin/gastrin-releasing peptide receptor and blocks the bombesin-stimulated increase in [Ca2+]i within the same concentration range that causes sustained activation of c-Jun N-terminal kinase and extracellular signal-regulated protein kinase (ERK). The activation of c-Jun N-terminal kinase by SpD and bombesin is blocked by dominant negative inhibition of G(alpha12). The ERK activation by SpD is pertussis toxin-sensitive in contrast to ERK activation by bombesin, which is pertussis toxin-insensitive but dependent on epidermal growth factor receptor phosphorylation. SpD does not simply act as a partial agonist but differentially modulates the activation of the G-proteins G(alpha12), G(i), and G(q) compared with bombesin. This unique ability allows the bombesin receptor to couple to G(i) and at the same time block receptor activation of G(q). Our results provide direct evidence that SpD is acting as a "biased agonist" and that this has physiological relevance in small cell lung cancer cells. This validation of the concept of biased agonism has important implications in the development of novel pharmacological agents to dissect receptor-mediated signal transduction and of highly selective drugs to treat human disease.
Topics: 3T3 Cells; Animals; Binding Sites; Blotting, Western; Bombesin; COS Cells; Calcium; Cell Line; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Inhibitors; GTP-Binding Proteins; Genes, Dominant; Humans; Ligands; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; Models, Theoretical; Pertussis Toxin; Phosphorylation; Protein Binding; Quinazolines; Rats; Receptors, Bombesin; Signal Transduction; Substance P; Time Factors; Tumor Cells, Cultured; Tyrphostins; Virulence Factors, Bordetella
PubMed: 11323408
DOI: 10.1074/jbc.M009772200 -
International Journal of Molecular... Apr 2015Bombesin receptor subtype-3 (BRS-3) is an orphan G-protein-coupled receptor (GPCR) member of the bombesin receptor family. Several studies have suggested an association...
Bombesin receptor subtype-3 (BRS-3) is an orphan G-protein-coupled receptor (GPCR) member of the bombesin receptor family. Several studies have suggested an association between obesity, alterations in glucose metabolism, diabetes and the BRS-3 receptor. In this study, we focused on patients simultaneously diagnosed with obesity and type 2 diabetes (OB/T2D). The analysis of BRS-3 expression in the skeletal muscle of these patients revealed a marked decrease in the expression of BRS-3 at the mRNA (23.6 ± 1.3-fold downregulation, p<0.0001) and protein level (49 ± 7% decrease, p<0.05) compared to the normal patients (no obesity and diabetes). Moreover, in cultured primary myocytes from patients with OB/T2D, the synthetic BRS-3 agonist, [D-Try6,β-Ala11,Phe13,Nle14]bombesin6-14, significantly increased the phosphorylation levels of mitogen-activated protein kinase (MAPK), p90RSK1, protein kinase B (PKB) and p70s6K. Specifically, the ligand at 10-11 M induced the maximal phosphorylation of MAPKs (p42, 159 ± 15% of the control; p44, 166 ± 11% of the control; p<0.0001) and p90RSK1 (148 ± 2% of the control, p<0.0001). The basal phosphorylation levels of all kinases were reduced (p<0.05) in the patients with OB/T2D compared to the normal patients. Furthermore, the BRS-3 agonist stimulated glucose transport, which was already detected at 10-12 M (133 ± 9% of the control), reached maximal levels at 10-11 M (160 ± 9%, p<0.0001) and was maintained at up to 10-8 M (overall mean, 153 ± 7%; p < 0.007). This effect was less promiment than that attained with 10-8 M insulin (202 ± 9%, p = 0.009). The effect of the agonist on glycogen synthase a activity achieved the maximum effect at 10-11 M (165 ± 16% of the control; p<0.0001), which did not differ from that observed with higher concentrations of the agonist. These results suggest that muscle cells isolated from patients with OB/T2D have extremely high sensitivity to the synthetic ligand, and the effects are particularly observed on MAPK and p90RSK1 phosphorylation, as well as glucose uptake. Moreover, our data indicate that BRS-3 may prove to be useful as a potential therapeutic target for the treatment of patients with OB/T2D.
Topics: Biological Transport; Diabetes Mellitus, Type 2; Enzyme Activation; Female; Gene Expression; Glucose; Glucose Transport Proteins, Facilitative; Glycated Hemoglobin; Glycogen Synthase; Humans; Male; Middle Aged; Muscle Cells; Obesity; Receptors, Bombesin; Signal Transduction
PubMed: 25653074
DOI: 10.3892/ijmm.2015.2090 -
Anesthesiology Aug 2019Although spinal opioids are safe and effective, pruritus is common and distressing. The authors previously demonstrated in mouse spinal cord that interactions between...
BACKGROUND
Although spinal opioids are safe and effective, pruritus is common and distressing. The authors previously demonstrated in mouse spinal cord that interactions between μ-opioid receptor isoform 1D and gastrin releasing peptide receptor mediate morphine-induced scratch. The C-terminal of 1D inhibits morphine-induced scratch without affecting analgesia. The authors hypothesize that human spinal cord also contains itch-specific μ-opioid receptor isoforms which interact with gastrin releasing peptide receptor.
METHODS
Reverse transcription polymerase chain reaction was performed on human spinal cord complimentary DNA from two human cadavers. Calcium responses to morphine (1 μM) were examined using calcium imaging microscopy on human cells (HEK293) coexpressing gastrin releasing peptide receptor and different human μ-opioid receptor isoforms. The authors assessed morphine-induced scratching behavior and thermal analgesia in mice following intrathecal injection of morphine (0.3 nmol) and a transactivator of transcription peptide designed from C-terminal sequences of 1Y isoform (0, 0.1, and 0.4 nmol).
RESULTS
The authors demonstrated 1Y expression in the spinal cord dorsal horn. Morphine administration evoked a calcium response (mean ± SD) (57 ± 13 nM) in cells coexpressing both gastrin releasing peptide receptor and the 1Y isomer. This was blocked by 10 μM naltrexone (0.7 ± 0.4 nM; P < 0.0001), 1 μM gastrin-releasing peptide receptor antagonist (3 ± 2 nM; P < 0.0001), or 200 μM 1Y-peptide (2 + 2 nM; P < 0.0001). In mice, 0.4 nmol 1Y-peptide significantly attenuated morphine-induced scratching behaviors (scratching bouts, vehicle vs. 1Y-peptide) (92 ± 31 vs. 38 ± 29; P = 0.011; n = 6 to 7 mice per group), without affecting morphine antinociception in warm water tail immersion test (% of maximum possible effect) (70 ± 21 vs. 67 ± 22; P = 0.80; n = 6 mice per group).
CONCLUSIONS
Human μ-opioid receptor 1Y isomer is a C-terminal splicing variant of Oprm1 gene identified in human spinal cord. Cross-talk between 1Y and gastrin releasing peptide receptor is required for mediating opioid-induced pruritus. Disrupting the cross talk may have implications for therapeutic uncoupling of desired analgesic effects from side effects of opioids.
Topics: Aged; Analgesics, Opioid; Animals; Behavior, Animal; Cadaver; Disease Models, Animal; Humans; Male; Mice; Mice, Inbred C57BL; Morphine; Naltrexone; Narcotic Antagonists; Pruritus; Receptors, Bombesin; Receptors, Opioid, mu; Spinal Cord
PubMed: 31314749
DOI: 10.1097/ALN.0000000000002776