D-chiro-inositol prevents memory deficits in the 3xTg mouse model of Alzheimer’s disease in a sex-dependent manner.

Subida por la tutora, a petición de Beatriz Pacheco Sánchez.Alzheimer’s disease (AD) is recognized as an age-related neurodegenerative disorder, characterized by the aggregation and deposition of amyloid-β (Aβ) in plaques and neurofibrillary tangles (NFTs) composed of aggregates of hyperphosphorylated tau, a microtubule-associated protein. Sporadic AD, the most prevalent form of the disease, has been proposed to be associated with alterations on insulin signaling. Insulin regulates a series of cognitive processes, such as learning and memory formation and emerging data demonstrate pivotal roles for brain insulin resistance and insulin deficiency as mediators of neurodegeneration, particularly in AD. The present study describes the effect of oral dietary administration of D-Chiro-inositol (DCI), an inositol used against insulin resistance, on the occurrence of the cognitive impairment of 3xTg-AD mice. This so called “triple‐ transgenic mice” model mimics many critical aspects of AD neuropathology. To this end, we analysed the effects of daily oral administration DCI 200 mg/Kg/day in male and female 3xTg and WT mice of 9 and 3 months of age. In addition to the evaluation of memory on the Novel Object Recognition, both the locomotion and time in the centre zone of an open field (OF), and the locomotion and anxiety-like behaviour in the elevated plus maze (EPM) test, were monitored. Our results showed that the exploration time of the new object was remarkably higher in the male 3xTg-DCI group when compared to those obtained from the 3xTg-vehicle and WT- vehicle groups at a middle and older age (3 and 9 months). When the center exploration time was analyzed in OPF test, three-way ANOVA showed an effect of the genotype in middle-aged animals. In the EPM test, only 9 months old 3xtg-DCI females were more prone to explore the open-arm, suggesting DCI decreases anxiety levels related to AD. In conclusion, our study suggest that DCI prevents and improves in a sex-dependent manner, the cognitive impairment associated to AD.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Reduction of adult hippocampal neurogenesis modifies brain functional connectivity and enhances cocaine-seeking in mice

Recently, adult hippocampal neurogenesis has been proposed as a putative neuroplastic mechanism involved in those behavioural processes. In this work, we studied the effect of the inhibition of adult hippocampal neurogenesis using the DNA alkylating agent temozolomide (TMZ), in cocaine-induced conditioned place preference (CPP) behaviour. In a first experiment, we investigated both CPP acquisition/expression and the functional brain circuits underlying CPP expression in control and neurogenesis-reduced conditions by analysing c-Fos immunoreactivity (c-Fos IR) in hippocampal and extrahippocampal addiction-related areas. A second experiment was designed to study the involvement of adult-born neurons in the extinction and cocaine-induced reinstatement of drug-seeking in the CPP model. We performed two independent studies where adult hippocampal neurogenesis was inhibited either before or after the CPP was acquired. Our results showed that TMZ treatment had no effect on the acquisition of the cocaine-induced CPP, but c-Fos IR associated to the test trial (CPP expression) revealed an increased activity in some of the analysed brain areas in the CPP-TMZ mice. Correlational and multivariate analysis revealed that, under normal conditions, the hippocampus showed widespread functional connectivity with other brain areas and strongly contributed to the functional brain network associated with CPP expression. However, mice with reduced neurogenesis showed an alternative brain circuit. The results of the second experiment revealed that mice acquiring the cocaine-induced CPP under neurogenesis-reduced conditions were delayed in extinguishing their drug seeking behaviour. However, when neurogenesis was inhibited after CPP acquisition, extinction was not affected but an enhanced long-term CPP retention was found, suggesting that the role of the adult-born neurons may differ depending on whether they are generated before or after drug-contextual associations are established. Importantly, cocaine-induced reinstatement of CPP behaviour was increased in the TMZ mice, regardless of the time of neurogenesis inhibition.Universidad de Málaga. Andalucía Tech, Campus de Excelencia Internacional. Spanish Ministry of Economy and Competitiveness (PSI2013-44901-P to L.J.S.; Subprograma RETICS Red de Trastornos Adictivos RD12/0028/0001, to F.R.F.). Author E.C-O. holds a ‘Sara Borrell’ research contract from the Spanish Carlos III Health Institute, Spanish Ministry of Economy and Competitiviness (grant number CD12/00455). Author D.L.G-M. holds a ‘FPU’ grant from the Spanish Ministry of Education, Culture and Sports (grant number FPU13/04819)

Persistent changes in exploration and hyperactivity coexist with cognitive impairment in mice withdrawn from chronic cocaine

Repeated cocaine exposure induces lasting neurobehavioral adaptations such as cognitive decline in animal models. However, persistent changes in spontaneous –unconditioned- motor and exploratory responses are scarcely reported. In this study, mice were administered with cocaine (20 mg/kg/day) or vehicle for 12 consecutive days. After 24 days of drug abstinence, a behavioral assessment was carried out in drug-free conditions and in unfamiliar environments (i.e. no cocaine-associated cues were presented). The cocaine-withdrawn mice showed cognitive deficits in spontaneous alternation behavior and place recognition memory. Importantly, they also displayed hyperlocomotion, increased rearing activity and altered exploratory patterns in different tasks. In the forced swimming test, they were more active (struggled/climbed more) when trying to escape from the water albeit showing normal immobility behavior. In conclusion, in addition to cognitive deficits, chronic cocaine in rodents may induce long-lasting alterations in exploratory activity and psychomotor activation that are triggered even in absence of drug-related stimuli.This study was funded by grants from the Spanish Ministry of Economy and Competitiveness (MINECO, Agencia Estatal de Investigación –AEI-) cofounded by the European Regional Development Fund-FEDER, UE- (PSI2015–73,156-JIN to E.C–O.; PSI2017–82604R to L.J.S.), RETICS Red de Trastornos Adictivos (ERDF-EU; RD16/0017/0001 to F.R.F.) and University of Málaga (B4: ‘Ayudas para Proyectos Puente’to E.C–O). Funding for open access charge: Universidad de Málaga /CBUA. Authors M.C.M-P., F. A-G. and S. G-R. hold predoctoral grants from the Spanish Ministry of Science, Innovation and Universities (FPU17/00276 to M.C.M-P.; PRE2018–085673 to F.A-G.; and FPU18/00941 to S.G-R.). Author D.L.G.M. holds a postdoctoral grant from University of Málaga (A.3. Plan Propio de Investigación y Transferencia Universidad de Málaga)

LH-21, a CB1 antagonist, reduces hepatotoxic damage produced by paracetamol overdose in a mice model.

Drug-induced liver injury (DILI) is one of the main causes of hepatic acute failure. Paracetamol can cause it when is ingested in excessive doses, leading to the depletion of the antioxidant mechanisms of the hepatocytes and a series of processes that conclude with cell death. One of the altered metabolic pathways is the endocannabinoid system (ECS), which has become a very interesting target to alleviate these events due to its involvement in inflammatory processes. For this reason, the triazole-derived compound LH-21, a cannabinoid receptor Cb1 antagonist, was used for the treatment of DILI in 8-week-old male C57BL/6 mice. In the present study, fasting mice were subjected to an oral overdose of paracetamol (300 mg/kg) and treated 2 hours later with 3 mg/kg of LH-21. After 24 hours, the animals were sacrificed and the livers were collected to determine the hepatic levels of metabolites related to antioxidant mechanisms, the expression of proteins involved in the generation of cellular damage and the transcription grade of the different components of the ECS. The observed results showed that LH-21 treatment raises GSH levels and total antioxidant capacity, in addition to reducing malondialdehyde values. Furthermore, the phosphorylation degree of Jnk and Stat3, as well as the activation status of Casp3, diminished. Regarding the ECS, the expression of Ppara, Cnr1, Cnr2 and Gpr55 did return to normal levels. This suggests that LH-21 effectively blocks the Cb1 activity, allowing the correct function of Ppar-α that promotes a cellular anti-inflammatory state and the relief of the symptoms produced by DILI. These results exhibit a promising perspective for the prevention or treatment of some toxic effects of paracetamol overdose with LH-21. Nevertheless, these findings are a first step to continue studying the involvement of the ECS in this type of liver disease and investigating the effectiveness of this Cb1 antagonist against the pathophysiology of DILI.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Enhanced markers of oxidative stress, altered antioxidants and NADPH-oxidase activation in brains from Fragile X mental retardation 1-deficient mice, a pathological model for Fragile X syndrome.

Política de acceso abierto tomada de: https://v2.sherpa.ac.uk/id/publication/6992Fragile X syndrome is the most common form of inherited mental retardation in humans. It originates from the loss of expression ofthe Fragile X mental retardation 1 (FMR1) gene, which results in the absence of the Fragile X mental retardation protein. However,the biochemical mechanisms involved in the pathological phenotype are mostly unknown. The availability of the FMR1-knockoutmouse model offers an excellent model system in which to study the biochemical alterations related to brain abnormalities in thesyndrome. We show for the first time that brains from Fmr1-knockout mice, a validated model for the syndrome, display higher levelsof reactive oxygen species, nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase activation, lipid peroxidation and proteinoxidation than brains from wild-type mice. Furthermore, the antioxidant system is deficient in Fmr1-knockout mice, as shown byaltered levels of components of the glutathione system. FMR1-knockout mice lacking Fragile X mental retardation protein werecompared with congenic FVB129 wild-type controls. Our results support the hypothesis that the lack of Fragile X mental retardationprotein function leads to a moderate increase of the oxidative stress status in the brain that may contribute to the pathophysiology ofthe Fragile X syndrome

Alpha-tocopherol protects against oxidative stress in the fragile X knockout mouse: an experimental therapeutic approach for the Fmr1 deficiency.

Política de acceso abierto tomada de: https://v2.sherpa.ac.uk/id/publication/4027Fragile X syndrome is the most common genetic cause of mental disability. The mechanisms underlying the pathogenesis remain unclear and specific treatments are still under development. Previous studies have proposed an abnormal hypothalamic–pituitary–adrenal axis and high cortisol levels are demonstrated in the fragile X patients. Additionally, we have previously described that NADPH-oxidase activation leads to oxidative stress in the brain, representing a pathological mechanism in the fragile X mouse model. Fmr1-knockout mice develop an altered free radical production, abnormal glutathione homeostasis, high lipid and protein oxidation, accompanied by stress-dependent behavioral abnormalities and pathological changes in the first months of postnatal life. Chronic pharmacological treatment with α-tocopherol reversed pathophysiological hallmarks including free radical overproduction, oxidative stress, Rac1 and α-PKC activation, macroorchidism, and also behavior and learning deficits. The restoration of the oxidative status in the fragile X mouse emerges as a new and promising approach for further therapeutic research in fragile X syndrome

Persistent drug-associated memories coexist with hippocampal-dependent cognitive decline and altered adult hippocampal neurogenesis in mice withdrawn from cocaine.

Aims: Using a new animal model (‘chronic’ cocaine-induced conditioned place preference –CPP- paradigm), this work studied whether the long-term maintenance of cocaine-associated memories was concomitant to cognitive impairment and adult hippocampal neurogenesis (AHN) alterations. Methods: Male c57BL/6J mice were submitted to a CPP task treated either with cocaine (20 mg/kg/day) or saline for 14 days (n=10 per group). Bromodeoxyuridine (BrdU) was administered to label the new hippocampal neurons generated one week after the last cocaine dose. After 28 drug-free days, mice were assessed for the CPP memory and on a battery of emotional and cognitive behavioral tests. After completion of behavior, brains were collected for AHN analysis. Results: In mice treated with cocaine, preference for the cocaine-paired compartment (CPP memory) persisted over time. In addition, the cocaine-withdrawn mice overall displayed normal emotional behavior but they showed hippocampal-dependent cognitive impairment for novelty recognition (object and place) and spatial (reference and working) memory. The number of BrdU+ cells was unaffected, suggesting that cocaine withdrawal did not impair basal AHN. However, the cocaine-withdrawn mice excessively increased the number immature hippocampal neurons (doublecortin+) after behavioral training, in direct correlation with their cognitive performance, probably as a result of effortful learning. Conclusions: The CPP memory induced by cocaine remains unaltered after a prolonged period of abstinence, accompanied by defective acquisition of new learnings. Since the doublecortin+ neurons correlated with better cognitive performance in the cocaine-withdrawn mice, strategies that increase AHN could alleviate neurocognitive deficits induced by cocaine.Plan Propio Universidad de Málaga Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Early maternal deprivation induces gender-dependent changes on the expression of hippocampal CB(1) and CB(2) cannabinoid receptors of neonatal rats.

Política de acceso abierto tomada de: https://v2.sherpa.ac.uk/id/publication/13858Early maternal deprivation (MD) in rats (24 h, postnatal day 9–10) is a model for neurodevelopmental stress. There are some data proving that MD affects the endocannabinoid system (ECS) in a gender-dependent manner, and that these changes may account for the proposed schizophrenia-like phenotype of MD rats. The impact of MD on cannabinoid receptor distribution in the hippocampus is unknown. The aim of this study is to evaluate the expression of CB1 and CB2 receptors in diverse relevant subregions (DG, CA1, and CA3) of the hippocampus in 13-day-old rats by immunohistochemistry and densitometry. MD induced a significant decrease in CB1 immunoreactivity (more marked in males than in females), which was mainly associated with fibers in the strata pyramidale and radiatum of CA1 and in the strata oriens, pyramidale, and radiatum of CA3. In contrast, MD males and females showed a significant increase in CB2 immunoreactivity in the three hippocampal areas analyzed that was detected in neuropil and puncta in the stratum oriens of CA1 and CA3, and in the polymorphic cell layer of the dentate gyrus. A marked sex dimorphism was observed in CA3, with females exhibiting higher CB1 immunoreactivity than males, and in dentate gyrus, with females exhibiting lower CB2 immunoreactivity than males. These results point to a clear association between developmental stress and dysregulation of the ECS. The present MD procedure may provide an interesting experimental model to further address the role of the ECS in neurodevelopmental mental illnesses such as schizophrenia. © 2008 Wiley-Liss, Inc.Consejería de Salud. Grant Number: PI-0220 Consejería de Innovación, Ciencia y Empresa. Grant Number: P05-CV1-1038 (Junta de Andalucía) Ministerio de Sanidad y Consumo. Grant Numbers: 2006/142, FIS 07/1226 Ministerio de Educación y Ciencia. Grant Number: SAF2006-07523 Red de trastornos adictivos. Grant Number: RD06/0001 Plan Nacional sobre Droga

Anxiety associated with palatable food withdrawal is reversed by the selective FAAH inhibitor PF-3845: A regional analysis of the contribution of endocannabinoid signaling machinery

Objective: Consumption of energy-dense palatable "comfort" food can alleviate stress and negative emotions, while abrupt withdrawal from a palatable diet can worsen these symptoms, causing difficulties with adherence to weight-loss diets. Currently, no pharmacological treatment is effective for obesity-related anxiety, so we investigated the endocannabinoid system (ECS), and specifically the fatty acid amide hydrolase (FAAH), as an interesting emerging target in this context because of its key role in the regulation of both energy homeostasis and emotional behavior. Methods: Rats were subjected to exposure and subsequent abstinence from a palatable cafeteria diet. During abstinence period, rats were treated with the selective FAAH inhibitor PF-3845 (10 mg/kg; intraperitoneal administration every other day). Results: Abstinent rats displayed an anxiogenic-like behavior and changes in the proteins of ECS signaling machinery in brain areas involved both in anxiety and food intake regulation. In particular, withdrawal caused a reduction of the expression of cannabinoid receptors in the nucleus accumbens and of enzymes diacylglycerol lipase alpha and monoacylglycerol lipase (MAGL) in the amygdala. Pharmacological inhibition of FAAH exerted an anxiolytic-like effect in abstinent animals and increased both MAGL expression in amygdala and CB2 expression in prefrontal cortex. Discussion: Overall, our results suggest that emotional disturbances associated with dieting are coupled with region-specific alterations in the cerebral expression of the ECS and that the enhancement of the endocannabinoid signaling by FAAH inhibition might represent a novel pharmacological strategy for the treatment of anxiety related to abstinence from palatable food.Funding for open access charge: Universidad de Málaga / CBUA European Regional DevelopmentFunds-European Union, Grant/Award Number:PI19/01577; Instituto de Salud Carlos III,Grant/Award Number: RETICS; Ministerio deCiencia e Innovaci on, Grant/Award Number:ERDF-EU RD16/0017/0001; Ministerodell'Università e della Ricerca, Grant/AwardNumber: 2012JTX3KL; PNRR-RomeTechnopole-FP

Overexpression of Cannabinoid CB2 Receptor in the Brain Induces Hyperglycaemia and a Lean Phenotype in Adult Mice

t is well known that the endocannabinoid system, through cannabinoid CB1 receptor activation,has an important role in the main aspects of energy balance (i.e. food intake, energy expenditureand glucose and fat metabolism), orchestrating all the machinery involved in body weight con-trol and energy homeostasis. A number of studies have revealed a crucial role of brain CB1receptors in these processes. However, functional cannabinoid CB2 receptors have also beendescribed in the brain, with no studies addressing their putative role in body weight control andglucose homeostasis. We have tested this hypothesis by analysing fasting-induced feeding, bodyweight, some hypothalamic neuropeptides, glucose tolerance and plasma hormones in an animalmodel specifically overexpressing CB2 receptors in the central nervous system. We found thatspecific overexpression of CB2 receptors in the brain promoted higher basal glucose levels,decreased fasting-induced feeding and, eventually, led to a lean phenotype and glucose intoler-ance. These findings could not be attributed to decreased locomotor activity, increased anxietyor depressive-like behaviours. The expression of relevant neuropeptides such as pro-opiomelano-cortin and galanin in the arcuate nucleus of the hypothalamus was altered but not those of theCB1 receptor. Indeed, no changes in CB1 expression were found in the liver, skeletal muscle andadipose tissue. However, cannabinoid CB1 and CB2 receptor expression in the endocrine pan-creas and glucagon plasma levels were decreased. No changes in plasma adiponectin, leptin,insulin and somatostatin were found. Taken together, these results suggest a role for centralcannabinoid CB2 receptors in body weight control and glucose homeostasis