Effects of galanin n-terminal fragment (1-15) in the light/dark and tail suspension tests

Galanin N-terminal fragment (1-15) [Gal(1-15)] is involved in mood regulation. The intracerebroventricular (icv) administration of Gal(1-15) produces a depressive-like behaviour in the forced swim test (FST) and an anxiety-like behaviour in the open field test (OF) in rats. In this work we analyze the effect of Gal(1-15) in two more behavioural tests, the tail suspension test (TLT) and the light/dark test. Gal(1-15) (3nmol), effective dose in FST and OF, or artificial cerebrospinal were injected in animals (n=5-8) 15 minutes before the test. Behavioural assessment were conducted with at least one week between tests. Student’s t-test was used for comparision between experimental groups. In the light/dark test we analyzed during 5 min three parametres as indicators of anxiety-like behaviour. Gal(1-15) significantly reduced the time spent in the light compartiment by 52% (p<0.05) and the latency time for entering the dark box by 65% (p<0.05). An increased in the latency time for re-entering the light box was also observed (p<0.05). In the TST, total immobility time was analyzed during 6 min test as parameter indicator of depressive-like behaviour. Gal(1-15) significantly increased rat immobility by 16% (p<0.05). Our results describe that Gal(1-15) exerts strong depressive- and anxiety-like effects in these tests, indicating a potential role of Gal(1-15) in mood disorders. These results may give the basis for the development of novel therapeutic drugs targeting Gal(1-15) for depression and anxiety.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Junta de Andalucia CVI6476 // Spanish Ministry of Economy and Competitiveness (PSI2013-44901-P to L.J.S). Author A.F-B. holds a ‘FPI’ grant from the Spanish Ministry of Economy and Competitiviness (grant number: BES-2014-068426)

Galanin n-terminal fragment (1-15) induces an anxiety- and depressive-like behaviours in the light/dark and tail suspension tests

Galanin N-terminal fragment (1-15) [Gal(1-15)] is involved in mood regulation. We have shown that intracerebroventricular (icv) administration of Gal(1-15) produces a depressive-like behaviour in the forced swim test (FST) and an anxiety-like behaviour in the open field test (OF) in rats. In this work we analyze the effect of Gal(1-15) in two more behavioural tests, the tail suspension test (TLT) and the light/dark test. In light/dark test we studied during 5 min the latency time for entering the dark box, time spent in the light compartiment, and the latency time for re-entering the light box as parameters indicators of anxiety-like behaviour. In TLT total immobility time was analyzed during 6 min test as parameter indicator of depressive-like behaviour. Groups of rats (n=5-8) were injected icv with Gal(1-15) 3nmol, a dose effective in FST and OF, or artificial cerebrospinal fluid 15 minutes before the test. Behavioural assessment were conducted with at least one week between tests. Student’s t-test was used for comparation between experimental groups. In the light/dark test Gal(1-15) 3nmol significantly reduced the time spent in the light compartiment by 52% (p<0.05) and the latency time for entering the dark box by 65% (p<0.05). An increased in the latency time for re-entering the light box was also observed (p<0.05). This pattern of results reflects an anxiogenic-like effects of Gal(1-15) in this test. In the TST, the administration of Gal(1-15) 3nmol significantly increased rat immobility by 16% (p<0.05) indicating a depressive-like effect. These results confirm the depressive- and anxiety-like effects of Gal(1-15) in rats. Future therapeutic strategies based on these results could be developed for depression and anxiety disorders. This work has been supported by the Junta de Andalucia CVI6476 and Spanish Ministry of Economy and Competitiveness (PSI2013-44901-P to L.J.S)Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. This work has been supported by the Junta de Andalucia CVI6476 and Spanish Ministry of Economy and Competitiveness (PSI2013-44901-P to L.J.S)Author A.F-B. holds a ‘FPI’ grant from the Spanish Ministry of Economy and Competitiviness (grant number BES-2014-068426)

Los derechos de los animales en las corridas de Toros desde la perspectiva constitucional

Artículo CientíficoEl presente artículo realiza un análisis con respecto a las decisiones judiciales de la Corte Constitucional, en relación con los espectáculos taurinos y la minoría que disfruta de la tauromaquia, permitiendo profundizar en una temática actual, que se contrapone a los derechos reconocidos para los animales en el ordenamiento jurídico colombiano contenidos en la Ley 84 de 1989, que ha sido modificada por la Ley 1774 de 2016.De igual forma, se orienta a resaltar la importancia de las decisiones constitucionales para la protección de las minorías taurinas, entendidas como un sector de la población susceptible de ser amparada por el marco jurídico constitucional, imponiendo un límite a la manifestación de los derechos de los animales que se consagran en el actual marco jurídico colombiano, exaltando la pertinencia del análisis planteado.Introducción 1. Un recuento de los derechos de los animales y los sistemas jurídicos en la historia de occidente. 2. La protección jurídica de los animales en Colombia 3. La auromaquia como manifestación cultural. 4. las decisiones de la Corte Constitucional colombiana en relación con las corridas de toros. Conclusiones. Referencias.PregradoAbogad

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Servicios Centrales de Apoyo a la Investigación (SCAI)

Desde mediados de la década de los noventa, la UMA cuenta con el SCAI como referente internacional en el equipamiento científico. Sus más de veinte laboratorios se dividen en cuatro grandes áreas científicas: análisis químico, ciencias de la vida, microscopía y protección radiológic

A Role For Galanin N-Terminal Fragment (1-15) In Anxiety And Depression in rats

Ponencia InvitadaGalanin (GAL) is involved in several functions including mood regulation. The GAL N-terminal fragment (1-15) [GAL(1-15)] also participates at central level and a differential role of GAL(1-15) compared with GAL has been proposed. In this work we have analysed if GAL(1-15) contributes to depression- and anxiety -related behaviours using the forced swimming test, tail suspension test, open field and light/dark test. We tested the involvement of the GAL receptor 2 (GALR2) in GAL(1-15) effects with the GAL receptor antagonist M871 and with an in vivo model of siRNA GALR2 knockdown rats. The proximity of GALR1 and GALR1 was also examined with the proximity ligation assay (PLA). GAL(1-15) induced strong depression-like and anxiogenic-like effects in all the tests. The involvement of the GALR2 was demonstrated with M871 and with the siRNA GALR2 knockdown rats. The PLA indicated the existence of GALR1-GALR2 heteroreceptor complexes in the dorsal hippocampus and especially in the dorsal raphe nucleus. Our results indicate that GAL(1-15) exerts strong depression-related and anxiogenic-like effects and may give the basis for the development of drugs targeting GALR1-GALR2 heteroreceptor complexes in the raphe-limbic system for the treatment of depression and anxiety. This study was supported by Junta de Andalucía CVI6476.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

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Galanin decreases NPYY1R internalization and β- Arrestin2 recruitment

We have recently described a Galanin receptor 2(GALR2) and Neuropeptide Y Y1 receptor(NPYY1R) interaction at behavioural, cellular and receptor levels through GALR2/NPYY1R heterodimers. The aim of this work was to study if GALR2 and NPYY1R costimulation modified NPYY1R internalization and β-Arrestin recruitment after in HEK293T cells. HEK293T cells were transfected with NPYY1REGFPor β-Arrestin2GFP2 cloned with standard molecular biology techniques employing PCR and fragment replacement strategies. NPYY1REGFP/GALR2 and NPYY1R/GALR2 with β- Arrestin2GFP2 HEK293T coexpressing cells were incubated with NPY 1μM and/or GAL1μM, at different times. Antagonist studies were performed 15 min prior to the addition of agonist with NPYY1R antagonist BIBP3226 10μM or GALR2 antagonist M871 10 μM. Timed-interval images of NPYY1REGFP or β-Arrestin2GFP2 endosomes in different cell groups were acquired using a confocal microscope following agonist addition. Percentage of internalization was determined by Leica software analysis of total membrane fluorescence compared to total internal compartment fluorescence at the various time points. We observed that addition of NPY induced a rapid decrease in the cell surface expression of NPYY1REGFP and a redistribution of β-Arrestin2GFP2. In fact, we observed a maximum of internalization of 80% three minutes after the NPY stimulation. However, combined treatment with GAL and NPY induced a delay in the internalization of NPYY1REGFP, with a maximum of internalization thirty minutes after the co-stimulation. Moreover, a delay in the β-Arrestin2GFP2 redistribution was observed. The specific GALR2 antagonist M871 abolished these delays in internalization of NPYY1REGFP and β-Arrestin2GFP2 redistribution, suggesting that this effect was mediated through the coactivation of GALR2 and NPYY1R. These results demonstrate that costimulation with GAL and NPY delays the internalization of  NPYY1REGFP by decreasing recruitment of β-Arrestin2GFP2 and probably could change intracellular signaling. This study was supported by Junta de Andalucia CVI6476.Junta de Andalucia CVI6476.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Galanin receptor 2 modifies neuropeptide Y Y1 receptor internalization and β-Arrestin recruitment

We have recently described a Galanin receptor 2(GALR2) and Neuropeptide Y Y1 receptor(NPYY1R) interaction at behavioural, cellular and receptor levels through GALR2/NPYY1R heterodimers. The aim of this work was to study if GALR2 and NPYY1R costimulation modified NPYY1R internalization and β-Arrestin recruitment after in HEK293T cells. HEK293T cells were transfected with NPYY1REGFPor β-Arrestin2GFP2 cloned with standard molecular biology techniques employing PCR and fragment replacement strategies. NPYY1REGFP/GALR2 and NPYY1R/GALR2 with β- Arrestin2GFP2 HEK293T coexpressing cells were incubated with NPY 1μM and/or GAL1μM, at different times. Antagonist studies were performed 15 min prior to the addition of agonist with NPYY1R antagonist BIBP3226 10μM or GALR2 antagonist M871 10 μM. Timed-interval images of NPYY1REGFP or β-Arrestin2GFP2 endosomes in different cell groups were acquired using a confocal microscope following agonist addition. Percentage of internalization was determined by Leica software analysis of total membrane fluorescence compared to total internal compartment fluorescence at the various time points. We observed that addition of NPY induced a rapid decrease in the cell surface expression of NPYY1REGFP and a redistribution of β-Arrestin2GFP2. In fact, we observed a maximum of internalization of 80% three minutes after the NPY stimulation. However, combined treatment with GAL and NPY induced a delay in the internalization of NPYY1REGFP, with a maximum of internalization thirty minutes after the co-stimulation. Moreover, a delay in the β-Arrestin2GFP2 redistribution was observed. The specific GALR2 antagonist M871 abolished these delays in internalization of NPYY1REGFP and β-Arrestin2GFP2 redistribution, suggesting that this effect was mediated through the coactivation of GALR2 and NPYY1R. These results demonstrate that costimulation with GAL and NPY delays the internalization of  NPYY1REGFP by decreasing recruitment of β-Arrestin2GFP2 and probably could change intracellular signaling. This study was supported by Junta de Andalucia CVI6476.Junta de Andalucia CVI6476.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Antidepressant-like effects induced by galanin 2/neuropeptide Y Y1 heterodimers in the dentate gyrus of the hippocampus

Previously, we have described the Galanin (GAL) and Neuropeptide Y Y1 (NPYY1) interactions through GAL receptor 2 and NPYY1 receptor 1 (GALR2/NPYY1R) heterodimers in the Dentate Gyrus (DG) of the Hippocampus, using autoradiographic, in situ hybridization and in situ proximity ligation assay(PLA) (1,2). The current work is to evaluate GALR2 and NPYY1R interactions in relation to depression-like behaviour and c-Fos expression in the Hippocampal DG. Rats (n=6-8) were forced to swim for a 15-min period (pre-test) and 24 h later were subjected to a 5-min swimming session (test) 15 min after the administration alone or in combination of GAL, the NPYY1R agonist [Leu31,Pro34]NPY and the GALR2 antagonist M871. The total duration of immobility, swimming, and climbing periods were scored during the test. For c-Fos immunohistochemistry, experimental groups of rats were anesthetized with sodium pentobarbital (100 mg/kg, i.p.) and perfused with 4 % Paraformaldehyde 90 min after icv injections. Then, brains were coronally sliced and immunostained. As primary antibodies, an antibody against c-Fos protein (1:5000, sc- 52, Santa Cruz Biotechnology, CA, USA), revealed with 3,3´-Diaminobenzidine (DAB) plus nickel, was used as an indirect marker of neural activity. The antibody to Calbindin-D28 k (1:1000, Santa Cruz Biotecnology, CA, USA), revealed with DAB, was used to outline the granular region since it marks mainly hippocampal granule cells. Sections were analyzed using the optical fractionator stereological method. We observed that icv injection of GAL and NPYY1R agonist significantly enhanced the decrease in the immobility (p<0,001) and the increase in the swimming behaviour (p<0,001) compared with the NPYY1R agonist alone. Moreover, a significant enhancement of the decrease in climbing behavior (p<0.05) was also observed. Furthermore, GALR2 is involved in this GALR/NPYY1R interaction, since the presence of the GALR2 antagonist M871 counteracted the enhancement of the decrease in immobility (p<0.01) and in climbing behavior (p<0.05) as well as the increase in swimming time (p<0.001) induced by the coadministration of GAL and NPYY1R agonist in the FST. Specific cells populations within DG subregions may be involved in this behavioural effect since the coadministration of GAL and NPYY1 agonist enhances the NPYY1R-mediated reduction (p<0.05) in the number of c-Fos immunoreactive nuclei in the polymorphic region. In this region, the GABA interneurons could be involved in the interaction since c-Fos IR colocalized with a GABAergic marker (GAD65/67) after NPYY1R agonist injection. Moreover, within the granular cells layer, GAL and NPYY1 agonist coadministration significantly increased c-Fos IR expression in the entire granular cell layer compared with GAL (p<0.05) and [Leu31,Pro34]NPY (p<0.01) alone. Again, the co-treatment with the GALR2 antagonist M871 completely reversed the GAL contribution to the responses in both regions, the polymorphic and the granular layer of the DG, demonstrating the involvement of GALR2 in the GAL actions. These results indicate that GALR2/NPYY1R interactions can provide a novel integrative mechanism in DG in depression-related behavior and may give the basis for the development of drugs targeting GALR2/NPYY1R heteroreceptor complexes in the DG of the hippocampus for the treatment of depression. Study supported by Junta de Andalucia CVI6476 and Proyecto Puente-Universidad de Málaga. 1. Galanin receptor 2-neuropeptide Y Y1 receptor interactions in the dentate gyrus are related with antidepressant-like effects. Narváez M, Borroto-Escuela DO, Millón C, Gago B, Flores-Burgess A, Santín L, Fuxe K, Narváez JA, Díaz-Cabiale Z. Brain Struct Funct 2016 Nov. 221(8):4129-4139. 2. Galanin receptor 2-neuropeptide Y Y1 receptor interactions in the amygdala lead to increased anxiolytic actions. Narváez M, Millón C, Borroto-Escuela D, Flores-Burgess A, Santín L, Parrado C, Gago B, Puigcerver A, Fuxe K, Narváez JA, Díaz-Cabiale Z. Brain Struct Funct. 2015 Jul;220(4):2289-301.Study supported by Junta de Andalucia CVI6476 and Proyecto Puente. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech