The Nociceptin/Orphanin FQ System and the Regulation of Memory

International audienceNociceptin/orphanin FQ (N/OFQ) is an endogenous neuropeptide of 17 amino acids, related to opioid peptides but with its own receptor, distinct from conventional opioid receptors, the ORL1 or NOP receptor. The NOP receptor is a G protein-coupled receptor which activates Gi/o proteins and thus induces an inhibition of neuronal activity. The peptide and its receptor are widely expressed in the central nervous system with a high density of receptors in regions involved in learning and memory. This review describes the consequences of the pharmacological manipulation of the N/OFQ system by NOP receptor ligands on learning processes and on the consolidation of various types of long-term memory. We also discuss the role of endogenous N/OFQ release in the modulation of learning and memory. Finally we propose several putative neuronal mechanisms taking place at the level of the hippocampus and amygdala and possibly underlying the behavioral amnestic or promnesic effects of NOP ligands

Neuropeptide FF/neuropeptide AF receptors (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

The Neuropeptide FF receptor family contains two subtypes, NPFF1 and NPFF2 (provisional nomenclature [10]), which exhibit high affinities for neuropeptide FF (NPFF, O15130) and RFamide related peptides (RFRP: precursor gene symbol NPVF, Q9HCQ7). NPFF1 is broadly distributed in the central nervous system with the highest levels found in the limbic system and the hypothalamus. NPFF2 is present in high density in the superficial layers of the mammalian spinal cord where it is involved in nociception and modulation of opioid functions

Effect of long-term exposure of SH-SY5Y cells to morphine: a whole cell proteomic analysis

BACKGROUND: Opiate addiction reflects plastic changes that endurably alter synaptic transmission within relevant neuronal circuits. The biochemical mechanisms of these adaptations remain largely unknown and proteomics-based approaches could lead to a broad characterization of the molecular events underlying adaptations to chronic drug exposure. RESULTS: Thus, we have started proteomic analyses of the effects of chronic morphine exposure in a recombinant human neuroblastoma SH-SY5Y clone that stably overexpresses the μ-opioid receptor. Cells were treated with morphine for 6, 24 and 72 hours, the proteins were separated by 2-D gel electrophoresis and stained with Coomassie blue, and the protein map was compared with that obtained from untreated cells. Spots showing a statistically significant variation were selected for identification using mass spectrometric analyses. CONCLUSION: A total of 45 proteins were identified, including proteins involved in cellular metabolism, cytoskeleton organization, vesicular trafficking, transcriptional and translational regulation, and cell signaling

Neuropeptide FF/neuropeptide AF receptors in GtoPdb v.2023.1

The Neuropeptide FF receptor family contains two subtypes, NPFF1 and NPFF2 (provisional nomenclature [12]), which exhibit high affinities for neuropeptide FF (NPFF, O15130) and RFamide related peptides (RFRP: precursor gene symbol NPVF, Q9HCQ7). NPFF1 is broadly distributed in the central nervous system with the highest levels found in the limbic system and the hypothalamus. NPFF2 is present in high density in the superficial layers of the mammalian spinal cord where it is involved in nociception and modulation of opioid functions

Direct Identification of a Peptide Binding Region in the Opioid Receptor-like 1 Receptor by Photoaffinity Labeling with [Bpa10,Tyr14]Nociceptin

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Denatured G-protein coupled receptors as immunogens to generate highly specific antibodies.

G-protein coupled receptors (GPCRs) play a major role in a number of physiological and pathological processes. Thus, GPCRs have become the most frequent targets for development of new therapeutic drugs. In this context, the availability of highly specific antibodies may be decisive to obtain reliable findings on localization, function and medical relevance of GPCRs. However, the rapid and easy generation of highly selective anti-GPCR antibodies is still a challenge. Herein, we report that highly specific antibodies suitable for detection of GPCRs in native and unfolded forms can be elicited by immunizing animals against purified full length denatured recombinant GPCRs. Contrasting with the currently admitted postulate, our study shows that an active and well-folded GPCR is not required for the production of specific anti-GPCR antibodies. This new immunizing strategy validated with three different human GPCR (μ-opioid, κ-opioid, neuropeptide FF2 receptors) might be generalized to other members of the GPCR family

Phosphoproteomic analysis of the mouse brain mu‐opioid (MOP) receptor

International audienceMany in vitro data have shown that the efficacy of several opioid drugs is correlated with differential mu‐opioid (MOP) receptor phosphorylation. Label‐free semiquantitative on‐line nanoflow liquid chromatography–tandem mass spectrometry (nanoLC–MS/MS) analyses were performed to compare the endogenous MOP receptor phosphorylation patterns of mice administered with morphine, etonitazene and fentanyl. The analysis identified S363, T370 and S375 as phosphorylated residues in the carboxy‐terminus. Only T370 and S375 were regulated by agonists, with a higher propensity to promote double phosphorylation for high efficacy agonists. Our study provides confirmation that differential agonist‐driven multi‐site phosphorylation of MOP receptor occurs in vivo and validate the use of MS to study endogenous GPCR phosphorylation

Proteomic analysis of immunostained, laser‐capture microdissected brain samples

International audienceAbstract Proteomic analysis is often performed on homogenized preparations of whole tissues, which does not provide any information about relevant biochemical changes in specific cell types. Laser‐capture microdissection (LCM) is a technique that is precise enough to dissect single cells within a tissue section. Phenotypically defined cells of interest may be visualized by immunostaining prior to microdissection. Previously published immunostaining protocols adapted to LCM require the use of very high antibody titers and very short incubation times. This raises the concern that low‐abundance antigens would not be detected and that antisera would be rapidly depleted. In addition, protein recovery from samples was not evaluated in most of these studies. Here, we describe an optimized immunostaining method based on immunofluorescence. By comparing two‐dimensional electrophoresis (2‐DE) results obtained from immunostained LCM brain tissue samples to those obtained from unstained, manually dissected samples, we demonstrated that immunofluorescent staining gave comparable protein recovery and similar resolution of protein spots on 2‐DE gels. Moreover, matrix‐assisted laser desorption/ionization‐time of flight (MALDI‐TOF) mass spectrometry analysis of selected spots from gels derived from control and immunostained LCM samples revealed that the immunostaining process had minimal effect on protein identification. LCM of immunofluorescently labeled tissue sections is a practical and powerful method to perform proteomic studies on specifically defined cell groups

Pharmacological insight into the activation of the human neuropeptide FF2 receptor

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