Cognitive reserve mediates the severity of certain neuropsychological deficits related to cocaine use disorder

Introduction: The concept of cognitive reserve (CR) is being considered in the field of substance use disorder (SUD) by observing that there are individuals whose brain alterations are not related to the cognitive symptomatology they present. Aims: Our aims were to characterise the possible neuropsychological deficits in a sample of subjects with SUD compared to a control group and to determine whether the degree of CR is a mediator in the cognitive functioning of these patients. Methods: To perform these objectives, the study involved a sample of subjects with SUD in outpatient treatment and a control group. A CR questionnaire and a comprehensive neuropsychological assessment were administered, and we also collected data related to drug consumption and psychological well-being. Results: The SUD group showed poorer performance compared to the control group in several cognitive domains (attention, declarative memory, executive functions and emotional perception), as well as in psychological comfort. Interestingly, we observed that the deficits found in attention, declarative memory and executive functions were mediated by the CR level of the participants, an effect that we did not observe in the rest of the variables registered. Conclusion: Our results suggest that long-term drug consumption leads to cognitive deficits and affects the psychological well-being of the subjects. Moreover, the CR should be taken into account during the assessment and rehabilitation of patients with SUD due to its protective role against certain neuropsychological deficits.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

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)

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

Influence of neurogenic improvement strategies on extinction and reinstatement of cocaine-induced conditioned place preference

AIMS: Modulation of adult hippocampal neurogenesis (AHN) has been shown to influence the maintenance of drug-context associations. We aimed to study whether the enhancement of AHN by using a water maze spatial learning task (WM), solely or under conditions of neurogenesis stimulation (forced treadmill exercise), could facilitate extinction and prevent primed reinstatement of cocaine-context associations. METHODS: Adult male C57BL/6J mice (N=37) were trained in the Conditioned Place Preference (CPP) paradigm with ascending doses of cocaine (2, 4, 8, 16 mg/kg/d) and subsequently received bromodeoxyuridine (BrdU) injections to label newborn neurons. Then, experimental groups were submitted to 12 days of scheduled exercise and/or 8 days of spatial training in the WM. Sedentary and/or untrained groups stayed undisturbed in their home cages. When BrdU+ cells reached maturation (~6 weeks-old), all mice were tested for CPP memory retrieval. Finally, animals were submitted to forced CPP extinction and tested for CPP extinction and cocaine-primed reinstatement. RESULTS: Animals submitted either to the scheduled exercise protocol, training in the WM or both strategies combined, required fewer sessions to extinct cocaine-CPP associations than control animals. Furthermore, animals submitted to both environmental strategies showed a reduced reinstatement when compared to sedentary animals. These effects are partially related to the functional integration of the newborn neurons in the hippocampus. CONCLUSIONS: Both environmental strategies, alone and combined, can reduce the long-term persistence of cocaine-context associations, being AHN associated with these beneficial effects.PSI2017-82604; PRE2018-085673; 08-2021-AREA3; B1-2020_06;Posdoc_21_00222; Posdoctoral_a32. I Plan Propio de la Universidad de Málaga. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

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

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

Behavioral traits predicting cocaine-conditioned place reference in mice: role of anxiety adn the basolateral amygdala

Aims. The individual susceptibility to cocaine addiction, a factor of interest in the understanding and prevention of this disorder, may be predicted by certain behavioral traits. However, these are not usually taken into account in research, making it difficult to identify whether they are a cause or a consequence of drug use. Methods. Male C57BL/6J mice underwent a battery of behavioral tests (elevated plus maze, hole-board, novelty preference in the Y maze, episodic-like object recognition memory and forced swimming test), followed by a cocaine-conditioned place preference (CPP) training to assess the reinforcing effect of the drug. In a second study, we aimed to determine the existence of neurobiological differences between the mice expressing high or low CPP by studying the number of neurons in certain addiction-related structures: the medial prefrontal cortex, the basolateral amygdala and the ventral tegmental area. Results. Anxiety-like behaviors in the elevated plus maze successfully predicted the cocaine-CPP behavior, so that the most anxious mice were also more likely to search for cocaine in a CPP paradigm. In addition, these mice exhibited an increased number of neurons in the basolateral amygdala, a key structure in emotional response including anxiety expression, without differences in the others regions analyzed. Conclusions. Our results suggest a relevant role of anxiety as a psychological risk factor for cocaine vulnerability, with the basolateral amygdala as potential common neural center for both anxiety and addiction.Universidad de Málaga, Campus de Excelencia Internacional Andalucía Tech. PSI2013-44901-P, FPU13/04819, CD12/00455, Red de Trastornos Adictivo

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

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