On the role of GPCR heteroreceptor complexes neuromodulation of the Claustrum

Esta conferencia invitada fue presentada dentro del simposium: Symposium S19 - Understanding the role of GPCR heteroreceptor complexes and their adaptor proteins in the neuronal networks of the brain in health and mental disordersG protein-coupled receptors (GPCRs) modulate the synaptic glutamate and GABA transmission of the claustrum. Our work focused on the transmitter-receptor relationships in the claustral catecholamine system and receptor-receptor interactions between kappa opioid receptors (KOR), dopamine receptor (D1R, D2R and D4R) and SomatostatinR2 (SSTR2) in claustrum. Methods used involved immunohistochemistry and in situ proximity ligation assay (PLA) using confocal microscopy. Double immunolabeling studies on D1R and tyrosine hydroxylase (TH) immunoreactivities (IR) demonstrated that D1R IR existed in almost all claustral and endopiriform nucleus nerve cell bodies, known as glutamate projection neurons, and D2R and D4R IR in large numbers of nerve cell bodies of the claustrum and endopiriform nucleus. However, only a low to moderate density of TH IR nerve terminals was observed in the endopiriform nucleus versus de few scattered TH IR terminals found in the claustrum. These results indicated that dopamine transmission in the rat operated via long distance dopamine volume transmission in the rat claustrum and endopiriform nucleus to modulate claustral-sensory cortical glutamate transmission. Large numbers of these glutamate projection neurons also expressed KOR and SSTR2 which formed KOR-SSTR2 together with D2R heteroreceptor complexes. The findings indicate that the sensory cortical glutamate drive on the glutamate claustral-cortical projection neurons is modulated by GPCRs and their receptor complexes located in the plasma membrane of these glutamate projection neurons. This can give the sensory cortical regions significant help in deciding on the salience to be given to various incoming sensory stimuli.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

A Transmembrane Single-Polypeptide-Chain (sc) Linker to Connect the Two G-Protein–Coupled Receptors in Tandem and the Design for an In Vivo Analysis of Their Allosteric Receptor- Receptor Interactions

A transmembrane (TM) single-polypeptide-chain (sc) linker can connect two G-protein–coupled receptors (GPCRs) in tandem. The priority of a gene-fusion strategy for any two class A GPCRs has been demonstrated. In the striatal function, dopamine (DA) plays a critical role. In the striatum, how the GPCR for adenosine, subtype A2A (A2AR), contributes to the DA neurotransmission in the “volume transmission”/dual-transmission model has been studied extensively. In addition to the fusion receptor, i.e., the prototype scA2AR/D2R complex (the GPCR for DA, subtype D2), several types were created and tested experimentally. To further elucidate this in vivo, we designed a new molecular tool, namely, the supermolecule scA2AR/D2R. Here, no experiments on its expression were done. However, the TM linker to connect the nonobligate dimer as the transient class A GPCR nanocluster that has not been identified at the cell surface membrane deserves discussion through scA2AR/D2R. Supramolecular designs, are experimentally testable and will be used to confirm in vivo the functions of the two GPCRs interactive in such a low specific signal to the nonspecific noise (S/N) ratio in the neurotransmission in the brain. The sc also has, at last, become straightforward in the field of GPCRs, similar to in the field of antibody

Galanin and neuropeptide y Y1 receptor agonist coinjection increases newborn cells proliferation on hippocampal dentate gyrus in rats

The hippocampus is a region in which neurogenesis persists throughout the lifespan in a wide variety of species including humans. Within the dentate gyrus of the hippocampus, the subgranular zone (SGZ) is maintained as a stem cell niche. We have previously shown that Galanin (GAL) interacts with Neuropeptide Y Y1 receptors (NPYY1R) in several regions of the central nervous system associated with mood and motivation. To examine the acute effects of GALR2/NPYY1R interactions on newborn cells proliferation we analyzed the effects of the intracerebroventricular (icv) of single injections with GAL and NPYY1 agonists or coadministered. Male Sprague-Dawley rats (n = 6-8 per group) were randomly assigned to the groups. Each group received i.c.v. injections of artificial Cerebro Spinal Fluid (aCSF), GAL or NPYY1R agonist [Leu31,Pro34]NPY alone or in combination. Intraperitoneal (ip) injections of exogenous cell DNA marker 5-bromo- 2-deoxyuridine (BrdU) 50mg/Kg were made at 2 and 4 hours after icv injections and 24 hours later rats were anesthetized, transcardially perfused and the brains collected for immunostaining to evaluate cell proliferation. Coadministration of GAL and NPYY1R agonist increased BrdU-labeled cells located in the SGZ (P<0,001) compared with aCSF, GAL and the NPYY1R-mediated hippocampal cell proliferation, These results will contribute to a better knowledge of the potential role of GAL and NPY family in mediating neurogenic actions and may give the basis for the therapeutic potential of targeting the GAL and NPY system in depressive disorders. Study supported by Proyecto Puente-Universidad de Málaga. Acknowledgements to Grupo Vithas.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Proyecto Puente-Universidad de Málaga

Potential of caveolae in the therapy of cardiovascular and neurological diseases.

Caveolae are membrane micro-domains enriched in cholesterol, sphingolipids and caveolins, which are transmembrane proteins with a hairpin-like structure. Caveolae participate in receptor-mediated trafficking of cell surface receptors and receptor-mediated signaling. Furthermore, caveolae participate in clathrin-independent endocytosis of membrane receptors. On the one hand, caveolins are involved in vascular and cardiac dysfunction. Also, neurological abnormalities in caveolin-1 knockout mice and a link between caveolin-1 gene haplotypes and neurodegenerative diseases have been reported. The aim of this article is to present the rationale for considering caveolae as potential targets in cardiovascular and neurological diseases

Galanin and Neuropeptide Y interaction leads to antidepressant effects through hippocampal cell proliferation.

Galanin (GAL) interacts with Neuropeptide Y Y1 receptors (NPYY1R) in several regions of the central nervous system associated with mood and motivation, through GAL receptor 2 and NPYY1 receptor 1 (GALR2/NPYY1R) heterodimers. The current work is to evaluate GALR2 and NPYY1R interactions concerning depression-like behavior and newborn cell proliferation in the ventral hippocampal Dentate Gyrus. Rats (n = 6-8 per group) were randomly assigned to the groups. Each group received i.c.v. injections of artificial Cerebro Spinal Fluid (aCSF), GAL or NPYY1R agonist [Leu31,Pro34]NPY alone or in combination and 24 h later rats were subjected to a 5-min swimming session (test). A different set of rats received ip injections of BrdU 50mg/Kg at 2 and 4 hours after icv injections. 24 hours later brains collected for immunostaining to evaluate cell proliferation. We observed that the icv injection of GAL and NPYY1R agonist significantly enhanced the decrease in the immobility and the increase in the swimming behavior compared with the NPYY1R agonist alone. Furthermore, GALR2 is involved in this GALR/NPYY1R interaction, since the presence of the GALR2 antagonist M871 counteracted all the parameters. In parallel, coadministration of GAL and NPYY1R agonist increased BrdU-labeled cells located in the SGZ compared with aCSF, GAL and the NPYY1R group. These results indicate that GALR2/NPYY1R interactions can provide a novel integrative mechanism in depression-related behavior and may give the basis for the development of drugs targeting GALR2/NPYY1R heteroreceptor complexes. Study supported by Proyecto UMA18-FEDERJA-100, Proyecto Puente-Universidad de Málaga, Proyecto Crowdfunding “Más Neuronas, menos depression” Universidad de Málaga. Acknowledgements to Catalina, Héctor Pittman Villarreal and Francisco Rodriguez LópezStudy supported by Proyecto UMA18-FEDERJA-100, Proyecto Puente-Universidad de Málaga, Proyecto Crowdfunding “Más Neuronas, menos depression” Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Acknowledgements to Catalina, Héctor Pittman Villarreal and Francisco Rodriguez Lópe

Improvement on hippocampal neuroproliferation through galanin and neuropeptide y y1 receptor agonist interaction

Neurogenesis persists in the hippocampus throughout the lifespan in a wide variety of species including humans. Within the dentate gyrus of the hippocampus, the subgranular zone (SGZ) is maintained as a stem cell niche. We have previously shown that Galanin (GAL) interacts with Neuropeptide Y Y1 receptors (NPYY1R) in several regions of the central nervous system associated with mood and motivation. To examine the acute effects of GALR2/NPYY1R interactions on newborn cells proliferation we analyzed the effects of the intracerebroventricular (icv) of single injections with GAL and NPYY1 agonists or coadministered. Male Sprague-Dawley rats (n = 6-8 per group) were randomly assigned to the groups. Each group received i.c.v. injections of artificial Cerebro Spinal Fluid (aCSF), GAL or NPYY1R agonist [Leu31,Pro34]NPY alone or in combination. Intraperitoneal (ip) injections of exogenous cell DNA marker 5-bromo-2-deoxyuridine (BrdU) 50mg/Kg were made at 2 and 4 hours after icv injections and 24 hours later rats were anesthetized, transcardially perfused and the brains collected for immunostaining to evaluate cell proliferation. Coadministration of GAL and NPYY1R agonist increased BrdU-labeled cells located in the SGZ (P<0,001) compared with aCSF, GAL and the NPYY1R-mediated hippocampal cell proliferation, These results will contribute to a better knowledge of the potential role of GAL and NPY family in mediating neurogenic actions and may give the basis for the therapeutic potential of targeting the GAL and NPY system in depressive disorders. Study supported by Proyecto Crowfunding “Más Neuronas, menos depression” Universidad de Málaga. Acknowledgements to Catalina, Héctor Pittman Villarreal and Francisco Rodriguez López.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Proyecto Crowfunding “Más Neuronas, menos depression” Universidad de Málaga. Acknowledgements to Catalina, Héctor Pittman Villareal and Francisco Rodriguez López

On the G-protein-coupled receptor heteromers and their allosteric receptor-receptor interactions in the central nervous system: focus on their role in pain modulation

The modulatory role of allosteric receptor-receptor interactions in the pain pathways of the Central Nervous System and the peripheral nociceptors has become of increasing interest. As integrators of nociceptive and antinociceptive wiring and volume transmission signals, with a major role for the opioid receptor heteromers, they likely have an important role in the pain circuits and may be involved in acupuncture. The delta opioid receptor (DOR) exerts an antagonistic allosteric influence on the mu opioid receptor (MOR) function in a MOR-DOR heteromer. This heteromer contributes to morphine-induced tolerance and dependence, since it becomes abundant and develops a reduced G-protein-coupling with reduced signaling mainly operating via beta-arrestin 2 upon chronic morphine treatment. A DOR antagonist causes a return of the Gi/o binding and coupling to the heteromer and the biological actions of morphine. The gender- and ovarian steroid-dependent recruitment of spinal cord MOR/kappa opioid receptor (KOR) heterodimers enhances antinociceptive functions and if impaired could contribute to chronic pain states in women. MOR1D heterodimerizes with gastrin-releasing peptide receptor (GRPR) in the spinal cord, mediating morphine induced itch. Other mechanism for the antinociceptive actions of acupuncture along meridians may be that it enhances the cross-desensitization of the TRPA1 (chemical nociceptor)-TRPV1 (capsaicin receptor) heteromeric channel complexes within the nociceptor terminals located along these meridians. Selective ionotropic cannabinoids may also produce cross-desensitization of the TRPA1-TRPV1 heteromeric nociceptor channels by being negative allosteric modulators of these channels leading to antinociception and antihyperalgesia

The zinc binding receptor GPR39 interacts with 5-HT1A and GalR(1) to form dynamic heteroreceptor complexes with signaling diversity

GPR39 is a class A G protein-coupled receptor involved in zinc binding and glucose homeostasis regulation, among other physiological processes. GPR39 was originally thought to be the receptor for obestatin peptide but this view has been challenged. However, activation of this receptor by zinc has been clearly established. Recent studies suggest that low GPR39 expression, due to deficient zinc levels, is involved in major depressive disorder. We have previously reported that zinc can alter receptor-receptor interactions and favor specific receptor interactions. In order to unravel the effect of zinc on specific G protein-coupled receptor association processes, we have performed FRET and co-immunopurification studies with GPR39 and 5-HT1A and GalR(1) which have been shown to dimerize. Our results suggest that zinc can modulate the formation of specific 5-HT1A-GPR39 and GalR(1)-5-HT1A-GPR39 heteroreceptor complexes under our experimental conditions.; We have analyzed the differences in signaling between the mono-homomeric receptors 5-HT1A, GalR(1) and GPR39 and the heteroreceptor complexes between them Our results show that the GPR39-5-HT1A heterocomplex has additive functionalities when compared to the monomeric-homomeric receptors upon receptor activation. In addition, the heterocomplex including also GalR(1) shows a different behavior, upon exposure to the same agonists. Furthermore, these processes appear to be regulated by zinc. These findings provide a rationale for the antidepressive effect widely described for zinc because pro-depressive heterocomplexes are predominant at low zinc concentration levels. (C) 2015 Elsevier B.V. All rights reserved.Peer ReviewedPostprint (author's final draft

Combined treatment with Sigma1R and A2AR agonists fails to inhibit cocaine self-administration despite causing strong antagonistic accumbal A2AR-D2R complex interactions: the potential role of astrocytes

Previous studies have indicated that acute treatment with the monoamine stabilizer OSU-6162 (5 mg/kg), which has a high affinity for Sigma1R, significantly increased the density of accumbal shell D2R-Sigma1R and A2AR-D2R heteroreceptor complexes following cocaine self-administration. Ex vivo studies using the A2AR agonist CGS21680 also suggested the existence of enhanced antagonistic accumbal A2AR-D2R allosteric interactions after treatment with OSU-6162 during cocaine self-administration. However, a 3-day treatment with OSU-6162 (5 mg/kg) failed to alter the behavioral effects of cocaine self-administration. To test these results and the relevance of OSU-6162 (2.5 mg/kg) and/or A2AR (0.05 mg/kg) agonist interactions, we administered low doses of receptor agonists during cocaine self-administration and assessed their neurochemical and behavioral effects. No effects were observed on cocaine self-administration; however, marked and highly significant increases using the proximity ligation assay (PLA) were induced by the co-treatment on the density of the A2AR-D2R heterocomplexes in the nucleus accumbens shell. Significant decreases in the affinity of the D2R high- and low-affinity agonist binding sites were also observed. Thus, in low doses, the highly significant neurochemical effects observed upon cotreatment with an A2AR agonist and a Sigma1R ligand on the A2AR-D2R heterocomplexes and their enhancement of allosteric inhibition of D2R high-affinity binding are not linked to the modulation of cocaine self-administration. The explanation may be related to an increased release of ATP and adenosine from astrocytes in the nucleus accumbens shell in cocaine self-administration. This can lead to increased activation of the A1R protomer in a putative A1R-A2AR-D2R complex that modulates glutamate release in the presynaptic glutamate synapse. We hypothesized that the integration of changes in presynaptic glutamate release and postjunctional heteroreceptor complex signaling, where D2R plays a key role, result in no changes in the firing of the GABA anti-reward neurons, resulting in no reduction in cocaine self-administration in the present experiments