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With a prevalence of 2-4% of the worldwide population, neurodevelopmental disorders (NDDs) comprise a heterogeneous group of disorders associated with neurodevelopmental dysfunction, including intellectual disability (ID), autism spectrum disorder (ASD), Down syndrome (DS) and attention-deficit/hyperactivity disorder (ADHD) among others. Last year, important steps in the mechanistic understanding of several NDDs have been achieved. New preclinical models demonstrated causality between PAK3 mutations and disorders associated with social deficiencies. ARID1B mutations have been linked to neuroectoderm specification in Coffin-Siris syndrome and DNA damage was established as an important pathologic mechanism in Aicardi- Gouti��res syndrome. Moreover, alterations in basic molecular processes including translation and histone acetylation have been established as major traits in the pathology of X-linked ID and Rett syndrome, revealing new pathogenetic mechanisms. Advances in bioinformatics have begun to shed light on the human repeatome, a largely unexplored part of our genome, and how alterations in these sequences have a central role in ASD. The role of mitochondria in neuropathology also advanced last year with the discovery of previously unknown vesicles derived from mitochondria, mitovesicles, with a putative role in ASD, DS and Alzheimer disease. An interesting discovery in the field is that during postnatal brain development, changes in genome architecture and transcriptional dynamics progress independently of sensory experience, a finding that will definitely impact the research in NDDs in the near future. Finally, our neurocentric views of NDDs is changing as new players such as astrocytes are revealed to be crucial in neuropathology. The role of astrocytes has started to be clarified for some pathologies such as ASD and DS, linking well-known genetic mutations to impaired astrocyte function., Free Neuropathology, Bd. 3 (2022)

OBJECTIVE Down syndrome (DS) is the most common form of chromosomal trisomy. Genetic factors in DS may increase the risk for diabetes. This study aimed to determine whether DS is associated with an increased incidence of diabetes and the relationship with obesity across the life span compared with control patients. RESEARCH DESIGN AND METHODS This matched population–based cohort study analyzed UK Clinical Practice Research Datalink data from 1990 to 2020. RESULTS A total of 9,917 patients with DS and 38,266 control patients were analyzed. Diabetes rates were higher in patients with DS (incidence rate ratio 3.67; 95% CI 2.43–5.55; P < 0.0001) and peaked at a younger age (median age at diagnosis 38 [interquartile range 28–49] years vs. 53 [43–61] years in control patients). Incidence rates (per 1,000 person-years) for type 1 diabetes mellitus were 0.44 (95% CI 0.31–0.61) in patients with DS vs. 0.13 (0.09–0.17) in control patients. Type 2 diabetes mellitus (T2DM) rates were higher in patients with DS versus control patients in age-groups from 5 years up to 34 years. In patients with DS, peak mean BMI was higher and at a younger age (males 31.2 kg/m2 at age 31 years; females 32.1 kg/m2 at 43 years) versus control patients (males 29.5 kg/m2 at 54 years; females 29.2 kg/m2 at 51 years). Obesity was associated with an increased incidence of T2DM. CONCLUSIONS At younger ages, the incidence of diabetes in patients with DS is up to four times that of control patients. Peak mean BMI is higher and established earlier in DS, contributing to T2DM risk. Further investigation into the relationship between obesity and diabetes in DS is required to inform treatment and prevention measures.

Trisomy of human chromosome 21 (HSA21) causes Down syndrome (DS). The trisomy does not simply result in the upregulation of HSA21--encoded genes but also leads to a genome-wide transcriptomic deregulation, which affect differently each tissue and cell type as a result of epigenetic mechanisms and protein-protein interactions. We performed a meta-analysis integrating the differential expression (DE) analyses of all publicly available transcriptomic datasets, both in human and mouse, comparing trisomic and euploid transcriptomes from different sources. We integrated all these data in a “DS network”. We found that genome wide deregulation as a consequence of trisomy 21 is not arbitrary, but involves deregulation of specific molecular cascades in which both HSA21 genes and HSA21 interactors are more consistently deregulated compared to other genes. In fact, gene deregulation happens in “clusters”, so that groups from 2 to 13 genes are found consistently deregulated. Most of these events of “co-deregulation” involve genes belonging to the same GO category, and genes associated with the same disease class. The most consistent changes are enriched in interferon related categories and neutrophil activation, reinforcing the concept that DS is an inflammatory disease. Our results also suggest that the impact of the trisomy might diverge in each tissue due to the different gene set deregulation, even though the triplicated genes are the same. Our original method to integrate transcriptomic data confirmed not only the importance of known genes, such as SOD1, but also detected new ones that could be extremely useful for generating or confirming hypotheses and supporting new putative therapeutic candidates. We created “metaDEA” an R package that uses our method to integrate every kind of transcriptomic data and therefore could be used with other complex disorders, such as cancer. We also created a user-friendly web application to query Ensembl gene IDs and retrieve all the information of their differential expression across the datasets., Author summary We analyzed all publicly available transcriptomic datasets on Down syndrome, the most common genetic intellectual disability, caused by trisomy of chromosome 21. Even though the triplicated genes are the same, the resulting transcriptional profile differs according to different tissues and states. Therefore, our goal was to understand which genes are most consistently deregulated both within and across different tissue macro-categories. We found that the effects of gene deregulation in Down syndrome originate from the upregulation of the triplicated genes but extend to others that are regulated or interact with the ones encoded on chromosome 21. This creates clusters of genes possibly involved in common biological or pathological processes, that are differentially expressed at the same time in different tissues or cellular states (co-DE expressed genes). The most consistent changes were specifically enriched in interferon related categories and neutrophil activation, reinforcing the concept that DS is an inflammatory disease. The genes that we detected can be very important for projects targeting key processes in Down syndrome for therapeutic purposes, as we show with the previously unreported upregulation of ETNPPL upon learning. Moreover, by applying our approach to different transcriptomic datasets, our method could be used for other complex disorders, such as cancer. This has been made easy thanks to development of “metaDEA”, an intuitive R package available on github.

In early development, the environment triggers mnemonic epigenomic programs resulting in memory and learning experiences to confer cognitive phenotypes into adulthood. To uncover how environmental stimulation impacts the epigenome and genome organization, we used the paradigm of environmental enrichment (EE) in young mice constantly receiving novel stimulation. We profiled epigenome and chromatin architecture in whole cortex and sorted neurons by deep-sequencing techniques. Specifically, we studied chromatin accessibility, gene and protein regulation, and 3D genome conformation, combined with predicted enhancer and chromatin interactions. We identified increased chromatin accessibility, transcription factor binding including CTCF-mediated insulation, differential occupancy of H3K36me3 and H3K79me2, and changes in transcriptional programs required for neuronal development. EE stimuli led to local genome re-organization by inducing increased contacts between chromosomes 7 and 17 (inter-chromosomal). Our findings support the notion that EE-induced learning and memory processes are directly associated with the epigenome and genome organization. We acknowledge support of the Spanish Ministry of Economy and Competitiveness (SAF2011-26216), “Centro de Excelencia Severo Ochoa 2017-2021,” SEV-2016-0571, the CERCA Programme/Generalitat de Catalunya and Jerome Lejeune Foundation, Swiss National Science Foundation Fellowship (PBLAP3_136878) and Co-funded by Marie Curie Actions to CNH. Resources for analyses conducted by SE-G were partially supported by the U.S. National Institutes of Mental Health Funds R01MH104341 and R01MH117790 and by the Social Sciences and Humanities Research Council of Canada (NFRFE-2018-01305). We acknowledge support of the Spanish Ministry of Science and Innovation to the EMBL partnership, Agencia Estatal de Investigaci n (PID2019-110755RB-I00/AEI / 10.13039/501100011033), the European Union’s Horizon 2020 Research and Innovation programme under grant agreement No 848077, Jerôme Lejeune Foundation, NIH (Grant Number: 1R01EB 028159-01), Marató TV3 (#2016/20-30). RP-R resources were supported by R01GM109215. We thank the support of the University of Tübingen for the Open Access Publication Funds contribution.

Epigallocatechin gallate (EGCG) is an inhibitor of DYRK1A, a serine/threonine kinase considered to be a major contributor of cognitive dysfunctions in Down syndrome (DS). Two clinical trials in adult patients with DS have shown the safety and efficacy to improve cognitive phenotypes using commercial green tea extract containing EGCG (45% content). In the present study, we performed a preclinical study using FontUp&reg, a new nutritional supplement with a chocolate taste specifically formulated for the nutritional needs of patients with DS and enriched with a standardized amount of EGCG in young mice overexpressing Dyrk1A (TgBACDyrk1A). This preparation is differential with previous one used, because its green tea extract has been purified to up 94% EGCG of total catechins. We analyzed the in vitro effect of green tea catechins not only for EGCG, but for others residually contained in FontUp&reg, on DYRK1A kinase activity. Like EGCG, epicatechin gallate was a noncompetitive inhibitor against ATP, molecular docking computations confirming these results. Oral FontUp&reg, normalized brain and plasma biomarkers deregulated in TgBACDyrk1A, without negative effect on liver and cardiac functions. We compared the bioavailability of EGCG in plasma and brain of mice and have demonstrated that EGCG had well crossed the blood-brain barrier.

Altres ajuts: This work was also supported by grants from the DCEXS is a 'Unidad de Excelencia María de Maeztu', funded by the AEI (CEX2018-000782-M) and Fundació La Marató de TV3 (20141210 to JF). Health conditions, immune dysfunction, and premature aging associated with trisomy 21 (Down syndrome, DS) may impact the clinical course of COVID-19. The T21RS COVID-19 Initiative launched an international survey for clinicians or caregivers on patients with COVID-19 and DS. Data collected between April and October 2020 (N=1046) were analysed and compared with the UK ISARIC4C survey of hospitalized COVID-19 patients with and without DS. The mean age of COVID-19 patients with DS in the T21RS survey was 29 years (SD = 18). Similar to the general population, the most frequent signs and symptoms of COVID-19 were fever, cough, and shortness of breath. Joint/muscle pain and vomiting or nausea were less frequent (p < 0.01), whereas altered consciousness/confusion were more frequent (p < 0.01). Risk factors for hospitalization and mortality were similar to the general population with the addition of congenital heart defects as a risk factor for hospitalization. Mortality rates showed a rapid increase from age 40 and were higher in patients with DS (T21RS DS versus non-DS patients: risk ratio (RR) = 3.5 (95%-CI=2.6;4.4), ISARIC4C DS versus non-DS patients: RR = 2.9 (95%-CI=2.1;3.8)) even after adjusting for known risk factors for COVID-19 mortality. Leading signs/symptoms of COVID-19 and risk factors for severe disease course are similar to the general population. However, individuals with DS present significantly higher rates of medical complications and mortality, especially from age 40. Down Syndrome Affiliates in Action, DSMIG-USA, GiGi's Playhouse, Jerome Lejeune Foundation, LuMind IDSC Foundation, The Matthew Foundation, NDSS, National Task Group on Intellectual Disabilities and Dementia Practices.

A major problem in treating obesity is the high rate of relapse to abnormal food-taking habits after maintaining an energy balanced diet. Alterations of eating behavior such as compulsive-like behavior and lack of self-control over food intake play a critical role in relapse. In this study, we used an operant paradigm of food-seeking behavior on two different diet-induced obesity models, a free-choice chocolate-mixture diet and a high-fat diet with face validity for a rapid development of obesity or for unhealthy food regularly consumed in our societies. A reduced operant performance and motivation for the hedonic value of palatable chocolate pellets was revealed in both obesity mouse models. However, only mice exposed to high-fat diet showed an increased compulsive-like behavior in the absence of the reinforcer further characterized by impaired operant learning, enhanced impulsivity and intensified inflexibility. We used principal component analysis to globally identify the specific behaviors responsible for the differences among diet groups. Learning impairment and inflexible behaviors contributed to a first principal component, explaining the largest proportion of the variance in the high-fat diet mice phenotype. Reinforcement, impulsion and compulsion were the main contributors to the second principal component explaining the differences in the chocolate-mixture mice behavioral phenotype. These behaviors were not exclusive of chocolate group because some high-fat individuals showed similar values on this component. These data indicate that extended access to hypercaloric diets differentially modifies operant behavior learning, behavioral flexibility, impulsive-like and compulsive-like behavior, and these effects were dependent on the exposure to each specific diet. This work was supported by the DG Research of the European Commission FP7 (#HEALTH-F2 2013-602891 and PHECOMP #LHSM-CT-2007-037669), the Spanish ‘RETICS-Instituto de Salud Carlos III’ (#RD16/0017/0020), the Spanish ‘Ministerio de Economia y Competitividad’ (#SAF-2014-59648P), the ‘Plan nacional sobre drogas’ (#PNSD-2013-5068), the Catalan Government ‘AGAUR-Generalitat de Catalunya’ (#2014-SGR-1547) and the Catalan foundation ‘La Marató de TV3’ (#2016/20-30). The laboratory of M.D. is supported by DIUE de la Generalitat de Catalunya (Grups consolidats SGR 2014/1125). This work was supported by Fondation Jérôme Lejeune (Paris, France), MINECO (SAF2013-49129-C2-1-R), CDTI (‘Smartfoods’) and EU (Era Net Neuron PCIN-2013-060). The CRG is a Center of Excellence Severo Ochoa SEV-2012-0208. The CIBER of Rare Diseases is an initiative of the ISCIII. The laboratory of C.N. acknowledges the funding from the Spanish Ministry of Economy and Finance (MINECO), grant number BFU2011-28575. J.E. receivedthe FI grant from Agència de Gestió d’Ajuts Universitaris i de Recerca (AGAUR) de la Generalitat de Catalunya

Supplemental material, JOP812103_supplemental_material for Metabolomics predicts the pharmacological profile of new psychoactive substances by Eulàlia Olesti, Ilario De Toma, Johannes G Ramaekers, Tibor M Brunt, Marcel·lí Carbó, Cristina Fernández-Avilés, Patricia Robledo, Magí Farré, Mara Dierssen, Óscar J Pozo and Rafael de la Torre in Journal of Psychopharmacology

Trabajo presentado a la X Reunión Científica Anual CIBERER, celebrada en San Lorenzo del Escorial del 23 al 24 de marzo de 2017.

The Barcelona Knowledge Hub of the Academia Europaea (AE- BKH)was set up in Barcelona in 2013 as the office for the Southern European region and the Mediterranean. The Academia Europaea is a pan-European, nongovernmental, not-for-profit association of over 4000 individual scien- tists and scholars who are recognized as experts and leaders in their own fields. It is committed to identifying topics of trans-European importance to science and scholarship, and provides, where appropriate, its expertise and its independent and impartial advice to European institutions, governments and international agencies concerning matters affecting science, scholarship and academic life in Europe. The AE-BKH organizes multidisciplinary activities that consider the perspective of the social sciences and the humanities, with scholarly aims as well as the goal of promoting the dissemination of science. One of the very special activities of the AE-BKH is the celebration of the pres- ent-day Disputatio of Barcelona as a remembrance of the original Disputatio of Barcelona held in 1263.Until present, four moder-day Disputationes have been held, from 2013 until 2016 [Contrib Sci 12:xx-xx (2016)]

La neurociencia requiere un escenario multidisciplinar con planteamientos y puntos de vista filosóficos, sociológicos, psiquiátricos, psicólogos, políticos, o biológicos: cada uno de ellos con una mirada complementaria y necesaria para abordar la relación entre mente, cerebro, y comportamiento humano. Ciertamente, los científicos no deben dejarse influir por objetivos de tipo político o cultural que nada tiene que ver con la esencia de nuestra profesión. La neurociencia nos provee de información que nos hace conscientes de nuestros sesgos subjetivos y nos permite estar alerta frente a nuestros miedos y a cómo éstos condicionan nuestra conducta y nuestra toma de decisiones. Es decir, nos permite luchar contra los estereotipos, y mejorar la aceptación social de la diversidad funcional.

a) Schematic representation of lentiviral vectors used to overexpress mmu-miR-155 and mmu-miR-802 (p-miR155 and p-miR802). b) Expression of mmu-miR155 and miR-802 in HeLa cells transfected with p-miR155 and p-miR802 (HeLa-miR-155-miR802). c) d2EGFP expression in HeLa and HeLa-miR-155-miR802 transduced with Lv-Control Lv-miR155T or Lv-miR155-802T. Top panel shows representative images of d2EGFP positive cells at 72-hours after transduction with 10 MOI of Lv-Control, LvmiR155Tor Lv-miR155-802T. Bottom panel shows the number of d2EGFP positive cells at different viraldoses. Image analysis was assessed using ImageJ software. Fig. S2. a) Principal component analysis of data obtained with the Agilent SurePrint G3 Mouse gene expression 8x60K Microarray (ID 028005). TheGEO accession number for our microarray data reported in this paper is GSE68074. Table of meandistances among the different genotypes and treatments. b) Prediction of miRNA targets based oncorrelation coefficients obtained from hippocampus treatment with sponge lentiviruses by different methods.Left panel shows the correlation prediction compared to the number of target sites predicted by TargetScan.Central panel shows the correlation prediction compared to the probability of conserved targeting(TargetScan PCT score). Right panel shows the correlation prediction compared to TargetScan contextscore. Fig. S3. a) Representation of Down’s syndrome gene expression dysregulation domanins (GEDDs)described by Letourneau and collaborators (black barplot overlay), and changes in expression in the miceorthologous genes observed in hippocampus upon the treatment with Lv-miR155-802T (grey barplotoverlay). Representation was performed using CIRCOS visualization package. b) Venn diagram of miRNApredicted targets on Lv-miR155-802T treated hippocampus based on the negative coefficients obtainedfrom miRComb analysis package for miR-155 (Corr_155 -) and miR-802 (Corr_802 -), genes that presenteda negative fold-change in between euploid and trisomic condition (TS/WT -) and genes that presented apositive fold-change in tisomic cells upon the XIST-induced HSA21 silencing (TS-XIST/TS +). (DOCX 3411 kb)

Obesity represents an important risk factor contributing to the global burden of disease. The current obesogenic environment with easy access to calorie-dense foods is fueling this obesity epidemic. However, how these foods contribute to the progression of feeding behavior changes that lead to overeating is not well understood and needs systematic assessment. Using novel automated methods for the high-throughput screening of behavior, we here examine mice meal pattern upon long-term exposure to a free-choice chocolate-mixture diet and a high-fat diet with face validity for a rapid development of obesity induced by unhealthy food regularly consumed in our societies. We identified rapid diet-specific behavioral changes after exposure to those high-caloric diets. Mice fed with high-fat chow, showed long-lasting meal pattern disturbances, which initiate with a stable loss of circadian feeding rhythmicity. Mice receiving a chocolate-mixture showed qualitatively similar changes, though less marked, consisting in a transient disruption of the feeding behavior and the circadian feeding rhytmicity. Strikingly, compulsive-like eating behavior is triggered immediately after exposure to both high-fat food and chocolate-mixture diet, well before any changes in body weight could be observed. We propose these changes as behavioral biomarkers of prodromal states of obesity that could allow early intervention. The laboratory of M.D. is supported by DIUE de la Generalitat de Catalunya (Grups consolidats SGR 2014/1125). This work was supported by Fondation Jérôme Lejeune (Paris, France), MINECO (SAF2013‐49129‐C2‐1‐R; SAF2016‐79956‐R), CDTI ('Smartfoods') and EU (Era Net Neuron PCIN‐2013‐060) and the Catalan foundation 'La Marató de TV3' (#2016/20‐30). The CRG is a Center of Excellence Severo Ochoa SEV‐2012‐0208. The CIBER of Rare Diseases is an initiative of the ISCIII. The laboratory of C.N. acknowledge the funding from the Spanish Ministry of Economy and Finance (MINECO), grant number BFU2011‐28575. J.E. received the FI grant from Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR) de la Generalitat de Catalunya. The laboratory of R.M. was supported by the DG Research of the European Commission FP7 (#HEALTH‐F2 2013‐602891 and PHECOMP #LHSM‐CT‐2007‐037669), the Spanish 'RETICS‐Instituto de Salud Carlos III' (#RD16/0017/0020), the Spanish 'Ministerio de Economia y Competitividad' (#SAF‐2014‐59648P), the 'Plan nacional sobre drogas' (#PNSD‐2013‐5068), the Catalan Government 'AGAUR‐Generalitat de Catalunya' (#2014‐SGR‐1547) and the Catalan foundation 'La Marató de TV3' (#2016/20‐30).

Background: People with Down syndrome (DS) are one of the highest risk groups for mortality associated with COVID-19, but outcomes may differ across countries due to different co-morbidity profiles, exposures, and societal practices, which could have implications for disease management. This study is designed to identify differences in clinical presentation, severity, and treatment of COVID-19 between India and several high-income countries (HICs). Methods: We used data from an international survey to examine the differences in disease manifestation and management for COVID-19 patients with DS from India vs HIC. De-identified survey data collected from April 2020 to August 2021 were analysed. Results: COVID-19 patients with DS from India were on average nine years younger than those from HICs. Comorbidities associated with a higher risk for severe COVID-19 were more frequent among the patients from India than from HICs. Hospitalizations were more frequent among patients from India as were COVID-19-related medical complications. Treatment strategies differed between India and HICs, with more frequent use of antibiotics in India. The average severity score of 3.31 was recorded for Indian DS in contrast to 2.3 for European and 2.04 for US cases. Conclusions: Presentation and outcomes of COVID-19 among individuals with DS were more severe for patients from India than for those from HIC. Global efforts should especially target vaccination campaigns and other risk-reducing interventions for individuals with DS from low-income countries. PH is supported by University Grant Commission, India (UGC/21/06/2015 (i)EU-V). SG is supported by Government of West Bengal (DST, WB) Grant No. SG/WBDST/S&T 1000114/2016 and UGC-UPEII, DST-PURSE programs (Grant No. UGC/859/UPE-2 BIO). AH is supported by the HERCULES Center (NIEHS P30ES019776) and the LuMind IDSC Foundation. The REDCap survey and database management system at Emory University was supported by Library Information Technology Services grant support (UL1 TR000424). ACSC is supported by the Alana USA Foundation, Awakening Angels Foundation, and the Infectious Diseases Society of America (IDSA). MD is supported by the Centre for Genomic Regulation Severo Ochoa excellence grant, the CIBER of Rare Diseases, DURSI 2017SGR595, and acknowledges the Spanish Ministry of Science, Innovation and Universities (MSIU) to the EMBL partnership, the Centro de Excelencia Severo Ochoa and CERCA (GenCat). AS is supported by the MRC (MR/ S011277/1; MR/S005145/1; MR/R024901/1), Lumind IDSC, The LeJeune Foundation and the European Commission (H2020 SC1 Gene overdosage and comorbidities during the early lifetime in Down syndrome GO-DS21- 848077). ML was supported by the National Institute for Health Research (NIHR) Biomedical Research Centre based at UCL Great Ormond Street Institute of Child Health/Great Ormond Street Hospital NHS Foundation Trust). DRdeA was partially supported by Spanish Fondo de Investigacion Sanitaria-Instituto Carlos III (FIS-ISCIII), grant No. PI19/00634; BAC has research support from Various Grateful Families of Patients. The Research Programme on Biomedical Informatics (GRIB) is a member of the Spanish National Bioinformatics Institute (INB), funded by ISCIII and EDER (PT17/0009/ 0014). The DCEXS is a ‘Unidad de Excelencia Marí a de Maeztu’, funded by the AEI (CEX2018-000782-M). The GRIB is also supported by the Agencia de G d ’AjutsUniversitarisi de Recerca (AGAUR), Generalitat de Catalunya (2017 SGR 00519). This study was also supported by the Fondo de InvestigacionesSanitario (FIS), Instituto de Salud Carlos III (PI18/00335 to MCI, PI14/ 01126 and PI17/01019 to JF), partly jointly funded by Fondo Europeo de Desarrollo Regional, Union Europea, Una manera de hacer Europa; the Jerome Lejeune Foundation (No.1319 Cycle 2019B to MCI); the National Institutes of Health (NIA grants 1R01AG056850 – 01A1; R21AG056974 and R01AG061566 to JF); Departament de Salut de la Generalitat de Catalunya, PlaEstrategic de RecercaiInnovaci o enSalut (SLT006/17/00119 to JF); and Fundacio La Marat o de TV3 (20141210 to JF).

Niemann-Pick disease type C (NPC) is a rare neurovisceral disease caused mainly by mutations in the NPC1 gene. This autosomal recessive lysosomal disorder is characterised by the defective lysosomal secretion of cholesterol and sphingolipids. No effective therapy exists for the disease. We previously described a deep intronic point mutation (c.1554-1009 G > A) in NPC1 that generated a pseudoexon, which could be corrected at the cellular level using antisense oligonucleotides. Here, we describe the generation of two mouse models bearing this mutation, one in homozygosity and the other in compound heterozygosity with the c.1920delG mutation. Both the homozygotes for the c.1554-1009 G > A mutation and the compound heterozygotes recapitulated the hallmarks of NPC. Lipid analysis revealed accumulation of cholesterol in the liver and sphingolipids in the brain, with both types of transgenic mice displaying tremor and ataxia at 7-8 weeks of age. Behavioural tests showed motor impairment, hyperactivity, reduced anxiety-like behaviour and impaired learning and memory performances, features consistent with those reported previously in NPC animal models and human patients. These mutant mice, the first NPC models bearing a pseudoexon-generating mutation, could be suitable for assessing the efficacy of specific splicing-targeted therapeutic strategies against NPC. This study was partly funded by grants from the Spanish Ministry of Science and Innovation (SAF2011-25431, SAF2013-49129-C2-1-R and SAF2014-56562-R) and from the Catalan Government (2014SGR/932 and SGR2014/1125), as well as the Addi and Cassi Fund. We are particularly grateful for the support from the Addi and Cassi Fund, who also provided the heterozygous mice. The Centre for Genomic Regulation has received support as a Severo Ochoa Centre of Excellence SEV- 2012-0208. We thank Alexandre Garcia for technical assistance. The authors are also grateful for the support from the Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), which is an initiative of the ISCIII. MGG was supported by a grant from the University of Barcelona (APIF).

The presence of an extra copy of Human Chromosome 21 (HSA21) in Down syndrome (DS) is one of the most complex genetic perturbations compatible with life and is the most common genetic cause of intellectual disability. Here, we performed the first comprehensive assessment of the cell-specific molecular changes that occur in the hippocampus of a mouse model of DS, Ts65Dn, by characterizing the transcriptome of tens of thousands of individual hippocampal neurons in parallel. This analysis revealed that the perturbations caused by the trisomy on the transcriptome are highly dependent on cellular identity, and led to the identification of genes, processes and cell composition alterations possibly involved in the DS pathogenetic mechanisms that were previously missed in bulk studies. Among them, we have unveiled the involvement of the long non-coding RNA gene Snhg11, specifically downregulated in the trisomic dentate gyrus, in adult neurogenesis and hippocampal-dependent cognitive phenotypes. We also identified a disruption of chromatin state that could contribute to the observed genome-wide transcriptional alterations and interfere with the patterns of gene expression required for memory formation. In particular, we found a marked decrease of the global histone acetylation levels in the trisomic hippocampus accompanied by a decrease of chromatin accessibility at gene promoters. Importantly, administering trisomic mice with a histone deacetylase inhibitor (HDACi) rescued histone hypoacetylation and restored expression of the memory-related Arc gene upon learning. In line with these results, the administration of HDACi also led to a total recovery of long-term recognition memory in trisomic mice. La trisomia del cromosoma humà 21 (HSA21), causa de la síndrome de Down (SD), és una de les alteracions genètiques més complexes compatibles amb la vida i és la causa genètica més comuna de la discapacitat intel·lectual. Per superar aquesta limitació, hem realitzat la primera avaluació exhaustiva dels canvis moleculars específics de cada tipus cel·lular que es produeixen a causa de la trisomia a l’hipocamp d’un model muri de la SD. Amb aquest objectiu, hem caracteritzat el transcriptoma de desenes de milers de neurones de l'hipocamp de ratolins wild-type i trisòmics. Aquesta anàlisi va revelar que les pertorbacions del transcriptoma causades per la trisomia depenen molt de la identitat de cada tipus cel·lular i ens han permès identificar gens, processos i alteracions de la composició cel·lular possiblement implicats en els mecanismes patogenètics de la SD que fins ara havien estat ignorats. Entre ells, hem descobert la implicació del long non-coding RNA Snhg11, que es troba reduït específicament al gir dentat dels ratolins trisòmics, a la neurogènesi adulta i als fenotips cognitius dependents de l'hipocamp. També hem identificat alteracions a l’estat de la cromatina que podrien contribuir a les alteracions transcripcionals observades a la SD i que podrien interferir amb els patrons d’expressió gènica que són necessaris per a la formació de la memòria. En particular, hem trobat una disminució significativa dels nivells globals d’acetilació de les histones a xviii l’hipocamp trisòmic, que és acompanyada per una disminució de l’accessibilitat de la cromatina als promotors dels gens. És important destacar que hem demostrat que l’administració d’un inhibidor de les deacetilases d’histones (HDACi) és suficient per recuperar aquesta manca d’acetilació i que també porta a una recuperació de l’expressió gènica associada a l’aprenentatge. D’acord amb aquests resultats, l’administració del HDACi també va comportar una recuperació total de la memòria de reconeixement a llarg termini als animals trisòmics tractats.

Tese de mestrado integrado em Engenharia Biomédica e Biofísica (Sinais e Imagens Médicas), Universidade de Lisboa, Faculdade de Ciências, 2020 Submitted by Cristina Manessiez (camanessiez@fc.ul.pt) on 2021-05-20T17:16:15Z No. of bitstreams: 1 ulfc126288_tm_Natalia_Marques.pdf: 5312686 bytes, checksum: cda648e8708aa9bbdc03125a6ee62eac (MD5) Made available in DSpace on 2021-05-20T17:16:27Z (GMT). No. of bitstreams: 1 ulfc126288_tm_Natalia_Marques.pdf: 5312686 bytes, checksum: cda648e8708aa9bbdc03125a6ee62eac (MD5) Previous issue date: 2020

Age-related hearing loss (ARHL) is the most common sensory deficit in the elderly. The disease has a multifactorial etiology with both environmental and genetic factors involved being largely unknown. SLC7A8/SLC3A2 heterodimer is a neutral amino acid exchanger. Here, we demonstrated that SLC7A8 is expressed in the mouse inner ear and that its ablation resulted in ARHL, due to the damage of different cochlear structures. These findings make SLC7A8 transporter a strong candidate for ARHL in humans. Thus, a screening of a cohort of ARHL patients and controls was carried out revealing several variants in SLC7A8, whose role was further investigated by in vitro functional studies. Significant decreases in SLC7A8 transport activity was detected for patient’s variants (p.Val302Ile, p.Arg418His, p.Thr402Met and p.Val460Glu) further supporting a causative role for SLC7A8 in ARHL. Moreover, our preliminary data suggest that a relevant proportion of ARHL cases could be explained by SLC7A8 mutations., eLife digest Age-related hearing loss affects about one in three individuals between the ages of 65 and 74. The first symptom is difficulty hearing high-pitched sounds like children’s voices. The disease starts gradually and worsens over time. Changes in the ear, the nerve that connects it to the brain, or the brain itself can cause hearing loss. Sometimes all three play a role. Genetics, exposure to noise, disease, and aging may all contribute. The condition is so complex it is difficult for scientists to pinpoint a primary suspect or develop treatments. Now, Guarch, Font-Llitjós et al. show that errors in a protein called SLC7A8 cause age-related hearing loss in mice and humans. The SLC7A8 protein acts like a door that allows amino acids – the building blocks of proteins – to enter or leave a cell. This door is blocked in mice lacking SLC7A8 and damage occurs in the part of their inner ear responsible for hearing. As a result, the animals lose their hearing. Next, Guarch, Font-Llitjós et al. scanned the genomes of 147 people from isolated villages in Italy for mutations in the gene for SLC7A8. The people also underwent hearing tests. Mutations in the gene for SLC7A8 that partially block the door and prevent the flow of amino acids were found in people with hearing loss. Some mutations in SLC7A8 that allow the door to stay open where found in people who could hear. The experiments suggest that certain mutations in the gene for SLC7A8 are likely an inherited cause of age-related hearing loss. It is possible that other proteins that control the flow of amino acids into or out of cells also may play a role in hearing. More studies are needed to see if it is possible to fix errors in the SLC7A8 protein to delay or restore the hearing loss.

During the last century the nervous system has been mainly studied from a reductionistic approach, based on the hypothesis that understanding in depth single neurons or limited neuronal populations would lead to general conclusions on brain function. However, to what extent anatomical details of single neurons can affect the wiring of the networks they form is a largely overlooked question. Intellectual disability provides an excellent opportunity to explore the relevance of fine structural details, because many disorders show specific architectural alterations that correlate with cognitive performance. In this Thesis, I aimed to study how the network topology of neuronal circuits is affected by dendritic architectural features in a mouse model of intellectual disability, namely Down's syndrome, and upon the rewiring effect of pro-cognitive treatment. I did so from three points of view: 1. The exploration of a 2D minimal computational model of cortical layer II/III parameterized by experimental data on dendritic tree architecture of healthy mice and two Down syndrome mouse models 2. The study of within-region morphological variations of hippocampal CA1 pyramidal neurons and their dependency of spatial embedding and cellularity in healthy mice and a Down syndrome mouse model. 3. The development of an experimental and computational framework for whole brain multiscale assessment and reconstruction. My work revealed that the dendritic tree architecture and the distribution of synaptic contacts have significant implications on how optimal single neurons are for information processing efficiency and storage capacity, and that those single-neuron features permeate to the network level, determining the computational capacities of neural ensembles. Also, I found position-dependent neuromorphological inhomogeneities in CA1 pyramids along with variations of neuronal cell density, suggesting that intrinsic properties of CA1 can vary across its extension. Those inhomogeneities were different in healthy and TgDyrk1A mice, possibly affecting emergent functional aspects. In my Thesis I faced challenges to bridge structure and function and to study morphological inhomogeneities at different scale (single cell and cell population). To solve xii those challenges, I developed computational methods for 3D mapping cellular population and dendritic density and assessed their validity. I also developed a computational modeling framework that allows the instantiation of multi-scale biologically realistic networks. Finally, I optimized the CLARITY whole-brain clearing technique and developed a pipeline to apply our population-based analysis and multi-scale modeling methods to the structural interrogation of whole brains, and to study the implications of the neuronal morphospace on the topology of neuronal circuitry. Durant l’últim segle, el sistema nerviós s’ha estudiat des d’un punt de vista reduccionista, basant-se en la hipòtesi que entendre en profunditat neurones individuals o fraccions petites de poblacions neuronals portaria a conclusions generals sobre la funció del cervell. De totes maneres, fins a quin punt detalls anatòmics de neurones individuals poden afectar la connectivitat de les xarxes que formen, és una qüestió que en gran part s’ha passat per alt. Les discapacitats intel·lectuals proporcionen una oportunitat excel·lent per explorar la rellevància de detalls estructurals, perquè molts trastorns cognitius mostren alteracions arquitectòniques específiques que correlacionen amb habilitats cognitives. En aquesta Tesi, pretenia estudiar com la topologia dels circuits neuronals és afectada per característiques arquitectòniques en un model murí de discapacitat intel·lectual, en concret de síndrome de Down, i per tractaments pro-cognitius amb efectes de remodel·lació de la xarxa. Ho he fet des de tres punts de vista: 1. L’exploració d’un model computacional 2D mínim de la capa cortical II/III parametritzat amb dades experimentals d’arquitectura dendrítica ens els nostres models de síndrome de Down. 2. L’estudi de neurones individuals, la seva diversitat i propietats morfològiques d’escala mesoscòpica en el model murí TgDyrk1A de síndrome de Down. 3. El desenvolupament d’un marc experimental i computacional per a l’estudi del problema des d’una perspectiva multi-escala. La meva feina ha mostrat que l’arquitectura dendrítica i la distribució de contactes sinàptics tenen implicacions significatives en l’optimalitat de neurones individuals per a l’eficiència en el processat d’informació i per a la capacitat d’emmagatzemar memòries, i que aquestes dues quantitats permeen al nivell de xarxa, determinant les capacitats computacionals de conjunts de neurones. També, he trobat variacions neuromorfològiques a CA1 dependents de la posició en neurones piramidals, acompanyades per variacions en densitat cel·lular, apuntat que propietats intrínseques de CA1 poden variar al llarg de la seva extensió. Aquestes inhomogeneitats eren diferents en ratolins sans i TgDyrk1A, possiblement tenint efectes en aspectes funcionals emergents concrets. xiv En la meva Tesi he afrontat reptes en lligar estructura i funció i en l’estudi de les inhomogeneïtats morfològiques en múltiples escales (de cèl·lula individual i de poblacions). Per a assolir aquests reptes, he desenvolupat mètodes computacionals per al mapejat 3D de poblacions cel·lulars i de densitats dendrítiques i he avaluat la seva validesa. També he desenvolupat un marc de modelització que permet l’instanciació multi-escala de xarxes neuronals biològicament realistes. Finalment, he optimitzat la tècnica de clarejat de cervell sencer CLARITY i he desenvolupat un pipeline per a aplicar les nostres eines d’anàlisi de poblacions i els mètodes multi-escala de model·lizatió per a l’anàlisi estructural de cervells sencers, i per a l’estudi de les implicacions del morfoespai neuronal en la topologia de la circuiteria neuronal.

Down syndrome (DS) is the most frequent genetic cause of intellectual disability. It is a multifaceted condition characterized by impairments in cognition, communication, behavior and/or motor skills resulting from abnormal brain development and function. Although DS is caused by the trisomy with more than 300 triplicated genes located on chromosome 21, there are a reduced number of dosagesensitive candidate genes that play a critical role in the pathogenesis of the disorder. Our group has made important contributions demonstrating that overexpression of a single gene, DYRK1A, is sufficient and necessary to recapitulate some of the DS phenotypes. Importantly, we could also demonstrate that the genetic, pharmacological or environmental normalization of its overdosage rescues behavioral, cognitive and neuronal phenotypes in preclinical studies with DS mouse models and in clinical trials in humans. In this Thesis I propose that the consequences of DYRK1A overexpression could spread along a complex intracellular network leading to a disease-causing network. For this reason, DYRK1A kinase normalization could be a good molecular target to restore the protein network functionality in DS. Here we have examined large-scale protein and phosphoprotein profiling to explore alterations caused by Dyrk1A overexpression and of pro-cognitive therapeutic strategies previously shown in the laboratory to normalize DYRK1A kinase activity and to promote cognition: i) environmental enrichment (EE), ii) and (-)- epigallocatechin-3-gallate (EGCG) and iii) the combination of both EGCG+EE. Using high-throughput tandem mass spectrometry (LC-MS/MS) in the hippocampus of TgDyrk1A mice we show that Dyrk1A overexpression in mice hippocampus primarily affects MAPK signaling and plasticity processes that could be related with impairments in recognition memory. Treatment with EGCG, EE, or their combination (EGCG+EE) restore some of the proteome and phosphoproteome alterations of Dyrk1A overexpression, possibly through the same mechanisms. La síndrome de Down (SD) és una de les causes mes freqüents de discapacitat intel·lectual. És una condició multifacètica caracteritzada per dèficits en cognició, comunicació, comportament i motors com a resultat d’un mal desenvolupament cerebral. Encara que la SD està causada per una trisomia que afecta a més de 300 gens del cromosoma 21, hi ha un reduït número de gens sensibles a dosis que juguen un paper crucial en la patogènesis d’aquesta patologia. El nostre grup de recerca ha fet contribucions importants en aquest camp i ha demostrat que la sobreexpressio d’un sol gen, DYRK1A, és suficient i necessària per recapitular alguns dels fenotips de la SD. També hem pogut demostrar que la normalització de la dosi de DYRK1A a nivells basals ja sigui de manera genètica, farmacològica, o a través d’una estimulació per enriquiment ambiental rescata alguns dels fenotips cognitius i neuronals en estudis preclinics amb models de ratolí però també amb humans. En aquesta tesi, es proposa que la sobreexpressió de DYRK1A condueix a canvis moleculars que s’expandeix a través d’una complexa xarxa interaccions intracel·lulars. Per aquesta raó, la normalització de DYRK1A podria ser una bona diana terapèutica per restaurar la funcionalitat de la xarxa proteica alterada en la SD. Hem examinat a gran escala el perfil proteòmic i fosfoproteòmic per tal d’explorar les alteracions causades per la sobreexpressió de Dyrk1A, així com les alteracions causades per les diferent estratègies que han sigut provades al laboratori que normalitzen la seva activitat quinasa: 1) enriquiment ambiental (EE), 2) (-)- epigallocatechin-3-gallate (EGCG) i 3) la combinació de EGCG+EE. Utilitzant espectrometria de masses per explorar les alteracions moleculars en l’hipocamp de ratolins que sobreexpressen Dyrk1A (TgDyrk1A) hem trobat alteracions que afecten la cascada de senyalització de les MAP quinases aixi com en processos relacionats amb la plasticitat que podrien estar relacionats amb els dèficits de reconeixement de memòria. Els tractaments amb EGCG, EE o la seva combinació han demostrat rescatar algunes de les alteracions proteòmiques i fosfoproteomiques induïdes per la sobreexpressió de Dyrk1A, possiblement mitjançant mecanismes comuns.

Down syndrome (DS) is a relatively common genetic condition caused by the triplication of human chromosome 21. No therapies currently exist for the rescue of neurocognitive impairment in DS. This review presents exciting findings showing that it is possible to restore brain development and cognitive performance in mouse models of DS with therapies that can also apply to humans. This knowledge provides a potential breakthrough for the prevention of intellectual disability in DS.

[eng] Obesity prevalence is higher in Down syndrome (DS) than in the general population. Beyond metabolic alterations, individuals with DS present increased impulsivity, a trait observed in obese people and in compulsive eaters that may affect their control of food intake. In this Thesis, we used a trisomic DS mouse model (Ts65Dn) to understand the behavioral component of obesity in DS and explore some possible underlying mechanisms. Our meal pattern analysis revealed longer and slowly meals in Ts65Dn mice, leading to reduced eating rate, which may be associated to the mandible hypoplasia described in both human and mice. When exposed to obesogenic environments, Ts65Dn mice showed higher preference for energy-dense food, gained more weight in specific conditions and scored higher in compulsivity and inflexibility tests than WT mice, as measured by binge eating during limited access and persistence of consumption of quine adulteration of energy-dense food. High performance liquid chromatography revealed reduced levels of dopamine in prefrontal cortex in Ts65Dn mice. This could lead to higher reward sensitivity that in turn would facilitate overeating as a compensatory response to restore optimal dopamine levels. Feeding behavior is also regulated by hormones and other circulating signals. We detected higher plasma leptin and glucose levels along with reduced insulin levels in Ts65Dn mice. Upon a glucose challenge, Ts65Dn mice showed reduced glucose-stimulated insulin response both in vivo and in vitro, suggesting a deficient insulin secretion or the reduced pancreatic mass. Indeed, we detected that Ts65Dn mice had altered plasma profile for some markers of inflammation and oxidative damage, in agreement with the high prevalence of autoimmune diseases and diabetes in DS people. We also explored the involvement of the serine/threonine kinase DYRK1A, a candidate DS gene, in obesity and feeding behavior. Dyrk1A overexpression was sufficient to recapitulate some behavioral aspects associated to overeating in DS, but with a distinct profile. We conclude that increased obesity prevalence in DS is explained by both metabolic and behavioral alterations, in part driven by a hypodopaminergic status, and that Dyrk1A overexpression is only involved in specific DS obesity phenotypes., [spa] La prevalencia de obesidad es más alta en el síndrome de Down (SD) que en la población general. Más allá de las alteraciones metabólicas, los individuos con SD tienen mayor impulsividad, rasgo común en personas obesas y en comedores compulsivos, que pueden afectar el control de la ingesta de alimentos. En esta Tesis, se ha utilizado un modelo de ratón trisómico (Ts65Dn) para comprender el componente de comportamiento en el desarrollo de la obesidad en SD. Nuestro análisis del patrón de ingesta mostró que los ratones Ts65Dn comen más lento que los euploides, lo que podría estar asociado con la hipoplasia mandibular descrita en ratones y humanos con SD. Cuando los ratones Ts65Dn son expuestos a ambientes obesogénicos, comen mayores cantidades de dietas hipercalóricas, engordan más en determinadas condiciones y puntúan más alto en pruebas de compulsividad e inflexibilidad que los ratones euploides. La cuantificación de los niveles de monoaminas mediante cromatografía líquida reveló que los ratones Ts65Dn presentan niveles más bajos de dopamina en corteza prefrontal. Dado que las dietas hipercalóricas promueven la liberación del neurotransmisor en el circuito de recompensa, el sobre consumo de las mismas podría indicar un intento de restaurar los niveles óptimos de dopamina. La regulación de la ingesta también depende de otras señales circulantes. Detectamos que los ratones Ts65Dn tienen mayores niveles de leptina y glucosa en plasma y niveles más bajos de insulina que los euploides. La administración exógena de glucosa produjo una menor respuesta secretoria de insulina en los ratones Ts65Dn in vivo e in vitro. Además, diversos marcadores de inflamación y estrés oxidativo son más elevados en los ratones Ts65Dn, en consonancia con la mayor incidencia de enfermedades autoinmunes y diabetes en personas con SD. En esta Tesis también se ha explorado la contribución de la proteína serina / treonina quinasa DYRK1A, un gen candidato para SD en la obesidad e ingesta. La sobreexpresión de Dyrk1A es suficiente para recapitular algunos comportamientos asociados a la ingesta compulsiva, pero con un perfil distinto al observado en el modelo trisómico. Concluimos que la prevalencia de la obesidad en SD se explica por alteraciones tanto metabólicas como conductuales, en parte como consecuencia de un estado de hipodopaminergia, y que la sobreexpresión de Dyrk1A está implicada en fenotipos específicos de la obesidad en SD.

Understanding the neural basis of conscious perception is still one of the central unsolved problems in neuroscience. Current theoretical models of conscious processing include the Global Workspace Theory, that proposes that for any stimulus to access to consciousness it should be broadcasted to different brain areas, making information, simultaneous and globally available among distant nodes in the network, through a network of neurons with long-range axons densely distributed in prefrontal, parieto-temporal, and cingulate cortices. Specifically, the hippocampus has been postulated as a critical hub, given its large structural and functional connectivity with distant brain areas. In this Thesis, I have taken advantage of the high spatial precision and signal-tonoise ratio afforded by intracranial recording in epileptic patients to explore signatures of this state of distributed ‘‘ignition’’ in two structures of the limbic system, the hippocampus and the amygdala. Electrophysiological data were recorded during the performance of two cognitive tasks for emotion recognition of faces (Emotion Recognition Test) and conscious perception of threatening words (Significant Words Paradigm). Specifically, I contrasted the fate of masked (subliminal) versus unmasked (conscious) words while recording from local sites in amygdala and hippocampus, using intracerebral depth electrodes in four epileptic patients. We explored two signatures of conscious processing: event related potentials (ERPs) and event related spectral perturbations (ERSPs) modifications discriminating emotion-related information (faces and words) and consciously perceived from subliminally processed stimuli. The analysis of the neuropsychological tests revealed a significant emotionspecific impairment in emotion recognition of faces, but no deficit was detected for perceiving emotion content of words in patients with epilepsy. ERP and ERSP analyses showed that amygdala, but not hippocampus has a role for emotion processing disregarding the quality of stimuli (faces or words); importantly, this was also observed in subliminal condition. On the other hand, when comparing conscious against subliminally perceived words, ERP and ERSP showed that amygdala and hippocampus are similarly involved in conscious processing; specifically, significant changes in voltage (ERP) and in theta and gamma frequencies (ERSP) were noted around 500 ms after target onset, supporting that conscious processing occurs between an early-perceptual window (0-200ms) and a later window in which more complex processing occurs (600-800ms). We conclude that those changes might constitute different measures of the same state of distributed ‘‘ignition’’ of a larger cortical network. Entender los mecanismos neuronales que dan lugar a la percepción consciente es todavía uno de los grandes problemas por resolver en neurociencia. Actualmente, los modelos teóricos sobre la consciencia incluyen la Teoría del Espacio de Trabajo Global, la cual propone que para que un estímulo acceda a la consciencia, éste debe ser transmitido a diferentes áreas cerebrales, de tal forma que su información esté disponible global y simultáneamente entre todos los nodos distantes que componen esa red, a través de una red de neuronas con axones de largo alcance densamente distribuidos entre las cortezas prefrontal, parietofrontal y cingulada. Específicamente el hipocampo ha sido propuesto como un nodo crítico, debido a sus numerosas conexiones estructurales y funcionales con regiones cerebrales distantes. En esta Tesis, he aprovechado la alta precisión espacial y relación señal-ruido que brindan los registros intracraneal en pacientes epilépticos, con el fin de explorar las firmas de este estado de “ignición” distribuida, desde dos estructuras del sistema límbico: el hipocampo y la amígdala, involucrados en el procesamiento de estímulos emocionales. Los datos electrofisiológicos fueron registrados durante la realización de dos tareas cognitivas de reconocimiento emocional de caras (Tarea de Reconocimiento de Emociones) y de palabras amenazantes (Paradigma de Palabras Significativas). Específicamente, contrasté el destino de registros de palabras enmascaradas (subliminales) versus no-enmascaradas (conscientes) mientras se registraban un sitiosl locales de amígdala e hipocampo en cuatro pacientes epilépticos. Exploramos las firmas del procesamiento consciente con: potenciales relacionados a evento (ERP) y perturbaciones espectrales relacionadas a evento (ERSP) en el hipocampo (theta) y en la amígdala (gamma) que discriminan entre estímulos emocionales conscientemente percibidos de aquellos procesados subliminalmente. El análisis de las pruebas neuropsicológicas reveló que los pacientes con epilepsia tienen una alteración significativa emoción-específica en el reconocimiento emocional de caras, pero ningún déficit en el reconocimiento de palabras. El análisis de ERP y ERSP mostró que la amígdala, pero no el hipocampo, tiene un papel en el procesamiento emocional independientemente de la calidad del estímulo (caras o palabras); importantemente, esto también fue observado en la condición subliminal. Por otro lado, cuando comparamos las palabras percibidas conscientemente contra aquellas percibidas subliminalmente, tanto los ERPS como los ERSP mostraron que la amígdala y el hipocampo están involucrados de manera similar en el procesamiento de la consciencia; específicamente, se observaron cambios significativos en el voltaje (ERP) y en frecuencias theta y gamma (ERSP) alrededor de 500ms después de la presentación del estímulo. Lo anterior apoya la idea que el procesamiento de la consciencia ocurre en una ventana temporal intermedia entre la percepción temprana (0-200ms) y un procesamiento cognitivo más complejo y tardío (600- 800ms). Concluimos que estos cambios pueden constituir diferentes medidas de un mismo estado de “ignición” distribuida de una red cortical más grande.

Clinical trials seeking to improve cognitive performance are in dire need of biochemical biomarkers that reflect the processes taking place in the brain. In Down syndrome, this information is crucial to understand the progression of cognitive decline, or to evaluate the beneficial effects of a treatment, but there are few available biomarkers for this condition. We have analyzed the association between biomarkers (biochemical, genetic), executive functions, and cognitive decline, in the context of a clinical trial, taking into account extrinsic factors (education, diet) that may obscure the interpretation of the obtained results. We found that altered lipid profile or increased homocysteine concentrations in plasma are associated to worse executive functions. Additionally, increased plasma concentrations of amyloid peptides are associated to early dementia signs. I also optimized a new technique of tissue and cell culture to obtain neuronal precursors from the nasal olfactory epithelium for biomarker studies. The results of this study should provide with new tools to evaluate treatment efficacy and cognitive decline risk in the context of clinical trials in Down syndrome., Els assajos clínics que cerquen la millora del rendiment cognitiu tenen la necessitat de disposar de biomarcadors que reflecteixin els processos que tenen lloc en el cervell. En la síndrome de Down aquesta informació és crucial per a entendre la progressió del declini cognitiu o per a avaluar els efectes beneficiosos d’un tractament però encara hi ha pocs biomarcadors disponibles per a aquesta afectació. Hem avaluat l’associació entre biomarcadors (bioquímics, genètics), funcions executives i declini cognitiu, en el context d’un assaig clínic, tenint en compte factors extrínsecs (educació, dieta) que poden afectar la interpretació dels resultats. Els nostres demostren que un perfil lipídic alterat o unes concentracions incrementades d’homocisteïna en plasma estan associats a pitjors funcions executives. També observem una associació entre concentracions incrementades de pèptids amiloides i signes primerencs de demència. També he optimitzat una nova tècnica de cultiu tissular i cel·lular per a obtenir precursors neuronals de l’epiteli olfactiu per a l’estudi de biomarcadors. Els resultats d’aquest estudi podrien proporcionar noves eines per a avaluar l’eficàcia de tractament i el risc de declini cognitiu en el context d’assaigs clínics en la síndrome de Down.

This Thesis aims at addressing the therapeutic potential of a neuroplasticity-targeted treatment for intellectual disability (ID) in Down syndrome (DS). The therapy consisted of the administration of the green tea catechin Epigallocatechin-3-gallate (EGCG), which is a natural modulator of Hsa21 candidate genes Dyrk1A and APP, in combination with cognitive stimulation. The assessment of the therapeutic efficacy of this intervention was performed using a translational approach, including preclinical studies with a mouse model of DS and clinical trials with humans with DS. In this work we show for the first time that this combined therapy significantly ameliorates cognitive deficits in mice and young adults with DS, by modifying brain neuronal networks structure and function., Esta Tesis tiene como objetivo examinar el potencial terapéutico de una intervención orientada a mejorar la neuroplasticidad cerebral, propuesta para mitigar la discapacidad intelectual (DI) en el síndrome de Down (SD). La terapia consistió en la administración de la catequina del té verde, Epigalocatequina-3-galato (EGCG), que es un modulador natural de dos genes candidatos que se encuentran en Hsa21, Dyrk1A y APP, en combinación con estimulación cognitiva. La evaluación de la eficacia terapéutica de esta intervención se realizó utilizando un enfoque de la translacional, incluyendo estudios preclínicos con un modelo de ratón de SD y ensayos clínicos con adultos jóvenes con SD. Este trabajo demuestra por primera vez que esta terapia combinada atenúa significativamente los déficits cognitivos en ratones y adultos jóvenes con SD, mediante la modificación de la estructura y función de redes neuronales en el cerebro.

New high-throughput behavioral systems enable the recording of continuous behavioral sequences with an unprecedented richness of signals and a deep temporal resolution. Automated systems offer neuroscience the opportunity to tackle in a new way the old question of how the brain orchestrates behavior and ultimately understand brain function itself, however, they accumulate large amounts of data leading to what is being termed Big Behavioral Data. The manipulation, analysis and contextualization of these data to obtain useful biological insights is not a trivial problem. This thesis presents Pergola, a computational framework to comprehensively analyze spontaneous longitudinal behaviors. Pergola provides access to a large set of mature genomic bioinformatics tools for the analysis and visualization of continuous behavioral recordings. I also explored multidimensional analysis techniques to help reducing the huge spatio-temporal dimensionality derived from behavioral recordings, and the high variability associated to all behavioral paradigms. This problem is addressed adapting Principal Component Analysis (PCA) for statistical inference on complex behaviors such as the recognition of learning strategies., Els nous sistemes d’alt rendiment per l’estudi del comportament permeten el enregistrement de senyals continues de comportament amb una riquesa de senyals i una resolució temporal sense precedents. Els sistemes automàtics ofereixen a la neurociència la oportunitat d’abordar d’una nova manera la vella qüestió de com el cervell orquestra el comportament i finalment entendre la pròpia funció cerebral, però a la vegada acumulen grans quantitats de dades, el que s’ha vingut a anomenar Big Behavioral Data. La manipulació, anàlisis i contextualització d’aquestes enormes quantitats de dates per a obtenir coneixements biològics útils no és un problema trivial. Aquesta tesi presenta Pergola, un marc computacional per analitzar exhaustivament els comportaments espontanis longitudinals. Pèrgola ofereix accés a un ampli conjunt d'eines madures de la bioinformàtica genòmica que poden ser usades per a l'anàlisi i visualització d'enregistraments contínues de comportament. També he explorat tècniques d'anàlisi multidimensionals per ajudar a reduir l'enorme dimensió espai-temporal derivada dels enregistraments de comportament, i l'alta variabilitat associada a tots els paradigmes de comportament. He adreçat aquest problema mitjançant l'Anàlisi de Components Principals (PCA) per la inferència estadística de comportaments complexos com per exemple, el reconeixement de les estratègies d'aprenentatge.

Estudios genéticos en pacientes sugieren que el gen del receptor tirosina quinasa de la neurotrofina de tipo 3 (NTRK3) podría contribuir en la patología del trastorno de pánico (PAND), un trastorno de ansiedad caracterizado por alteración y exageración del miedo. En esta tesis, proponemos que PAND podría estar relacionado con procesos anormales de aprendizaje asociativo, subyacentes a una alteración en la regulación del funcionamiento del circuito del miedo hipocampo - amígdala – corteza prefrontal media. Hemos estudiado esta hipótesis utilizando el único modelo genético de ratón validado de PAND, el TgNTRK3, que sobreexpresa el gen humano NTRK3 codificante para el receptor TrkC. Hemos demostrado que la sobreexpresión de NTRK3 en ratones produce: i) un incremento en la memoria de miedo dependiente de hipocampo que es además resistente a extinción; ii) la activación alterada del circuito del miedo durante el aprendizaje y almacenamiento de experiencias emocionales intensas negativas. Nuestros resultados sugieren que los mecanismos responsables están relacionados con una sobre excitabilidad en el hipocampo. Nuestro estudio confirma el papel del gen NTRK3 en la memoria de miedo patológico y sugiere posibles estrategias terapéuticas para bloquear el miedo exagerado en PAND dirigidas al sistema de excitación / inhibición y a la vía de señalización NT3-TrkC., The neurotrophin tyrosine kinase receptor type 3 (NTRK3) gene has been proposed to contribute to the pathological phenotype of panic disorder (PAND), an anxiety disorder characterized by perturbed and exaggerated fear. In this thesis we hypothesized that PAND could be related to abnormal associative fear learning processes, underlined by a deregulated functioning of the hippocampus – amygdala – medial prefrontal cortex fear circuit. We addressed this hypothesis, by using the unique validated genetic mouse model of PAND, the TgNTRK3, overexpressing the human NTRK3 gene, encoding for TrkC. We found that overexpression of NTRK3 in mice leads to: i) enhanced and extinction resistant hippocampal-dependent fear memories; ii) an aberrant activation of the fear circuit during learning and storage of fear-related information. The underlying mechanisms are possibly related to hippocampal overexcitability. Our study confirmed the role of the NTRK3 gene on pathological fear memories and suggests potential effective therapeutical strategies targeting the excitatory/ inhibitory system and the NT3-TrkC pathway to block exaggerated fear in PAND.

Esta Tesis Doctoral ha consistido en el estudio de los mecanismos implicados en la comorbilidad entre el abuso de drogas y los trastornos de ansiedad, así como en las regiones cerebrales responsables de la coexistencia de estos dos trastornos en modelos transgénicos. Nos hemos centrado en dos grandes familias de genes, que podrían actuar como interfaz genético en la comorbilidad entre trastornos de ansiedad y de abuso de sustancias: neurotrofinas por su intervención en el neurodesarrollo y en la neuroplasticidad, y elementos de sistemas de neurotransmisión concretos como son los receptores nicotínicos, cuya implicación en adicción y ansiedad ha sido previamente demostrada a través de estudios farmacológicos.1) HIPÓTESIS:La comorbilidad entre trastornos de ansiedad y de abuso de sustancias es un hecho bien establecido y que tiene importantes implicaciones clínicas, terapéuticas y pronósticas. Si bien los mecanismos patogenéticos de cada uno de estos trastornos por separado se conocen relativamente bien, el mecanismo que subyace a la comorbilidad no está bien delimitado. Se ha propuesto la existencia de factores genéticos, neurobiológicos y/o ambientales comunes a ambos trastornos, pero todavía no está claro qué genes podrían estar implicados o ser predisponenentes a esta comorbilidad. Nosotros proponemos que los factores genéticos de susceptibilidad para la patología dual deberían modular componentes de desarrollo, y componentes funcionales, que están implicados en la coexistencia entre ambos trastornos. Por esta razón el trabajo, se centra en genes de dos familias: el receptor de la neurotrofina-3 (NTRK3) y un cluster que contiene los genes de las subunidades, α3, α5, β4 de receptores nicotínicos (CHRNA3/A5/B4). Así, NTRK3 podría participar en el desarrollo y los mecanismos adaptativos a través de su implicación en proliferación, diferenciación y plasticidad neuronal, mientras que el cluster CHRNA3/A5/B4 podría estar implicado en la activación de circuitos neuronales específicos que podrían ser comunes a ambos trastornos.2) OBJETIVOS:El objetivo general de esta tesis ha consistido en el estudio de la posible implicación funcional de los genes objeto de estudio en elementos de la comorbilidad entre trastornos de ansiedad y abuso de sustancias mediante el uso de ratones transgénicos como modelo de aproximación experimental. Para ello hemos caracterizado el impacto de la sobreexpresión in vivo de los genes NTRK3 y CHRNA3/A5/B4 sobre el fenotipo conductual, neuromorfológico y neurofisiológico.Dado que intentamos comprender los mecanismos implicados en la comorbilidad entre el abuso de drogas y los trastornos de ansiedad, pero también delimitar las regiones cerebrales responsables de la co-ocurrencia de estos dos trastornos se han realizado estudios conductuales, farmacológicos con drogas de abuso como morfina o nicotina, y experimentos de caracterización histológica y molecular.3) CONCLUSIONES:Como conclusión general, los resultados obtenidos demuestran que alteraciones en los niveles de expresión de genes tan diferentes como los factores neurotróficos o los receptores nicotínicos, pueden modificar la actividad de regiones cerebrales implicadas en la aparición simultánea de diferentes trastornos psiquiátricos. De nuestros experimentos se desprende que el sistema mesocorticolímbico, junto con regiones troncoencefálicas como el LC, jugaría un papel clave en la comorbilidad de trastornos como la ansiedad, la adicción o la epilepsia. Un mecanismo de convergencia podría depender de los cambios en receptores glutamatérgicos, que son elementos clave en la plasticidad neuronal. Nuestros resultados apoyan la naturaleza poligénica de la patología dual y sugieren que los cambios de dosis de determinados genes podrían ejercer un papel esencial en la morbilidad de la misma., This Doctoral Thesis has been focused on understanding the mechanisms and the cerebral regions implicated in the commorbidity between drug abuse and anxiety disorders. To this aim we have focused on two families of genes that may act as genetic interface to the commorbidity between these neuropsychiatric disorders: 1/ neurotrophins, that may interfere in neurodevelopment and neuroplasticity, and 2/ elements of specific neurotransmitter systems such as nicotinic receptors, that have been implicated in addiction and anxiety, previously demonstrated by pharmacological studies. Our hypothesis proposes the existence of a development component and a functional component in the generation of an addictive and anxious brain. To this study we used two mouse models: 1/ A transgenic mouse model overexpressing the neurotrophin-3 receptor, TrkC (TgNTRK3), since it has been implicated in plasticity, proliferation and cellular differentiation with a possible pathogenetic intervention in the dual pathology, such as the noradrenergic system, and 2/ A transgenic mouse model overexpressing the α3, α5, β4 nicotinic receptor subunits (TgCHRNA3/A5/B4), candidates to the development of psychiatric disorders and addiction. To determine the gene overexpression effects on the appearance of this dual pathology we have performed pharmacological studies using drug abuse, such as nicotine and morphine. The results obtained suggest that alterations in the expression levels of these genes could be modifying the activity of different cerebral regions responsible for the commorbidity between both disorders in transgenic mouse models. Thus, our experiments suggest that the mesocorticolimbic system, as well brainstem regions, such as the Locus coeruleus, would play a key role in the commorbidity between anxiety and addiction. That could be due to alterations on glutamatergic receptors, key elements in neuronal plasticity.

The correct development and function of CNS is critical for brain health of the organism. Early or chronic stress causes prominent alterations in brain function, and affects the expression of neurotrophic factors in limbic brain regions involved in the regulation of mood and cognition. Recent evidences have opened the idea that in complex organisms, an altered expression of certain neurotrophins by stress could be involved in the onset and pathophysiology of most psychiatric disorders, such as depression, squizophrenia or anxiety disorders. It is hypothesized that altered levels of neurotrophic factors could contribute to the atrophy and cell death of these regions, including the hippocampus and prefrontal cortex, which would produce a malfunction in limbic-related areas, and as a consequence, a precipitation or worsening of psychiatric illnesses. We were interested in panic disorder pathophysiology, which is a stress-related disorder and is characterized by an altered cognitive processing of emotional information. Although little evidence has been found supporting a neurotrophic role in PD, recent data has revealed that NT-3/TrkC signaling might play a key role in limbic system morphology and function. Therefore, we suggest that NT-3/TrkC system is involved in PD pathogenesis. The main objective in the work of this doctoral thesis lie to determine the role of NTRK3 gene, that codifies for TrKC, in emotional cognition and stress response processes that underlies PD. To this end, we used a genetically modified mouse model of NTRK3 overexpression, which was validated as a model of PD. Here, it is characterized the effects produced by the increase of NTRK3 expression in the CNS, focusing in neural alterations that might influence changes in cognitive processes involved in coping strategies. Moreover, it is studied the mechanisms that underlie in these processes by different approaches, 1/physiologically, measuring the HPA axis response, 2/brain activation, analyzing the activation pattern to a stress stimulus, 3/cellular and gene expression profiling, characterizing key brain regions in cognitive processes, and 4/pharmacologically, studying neurotransmitters function.

El síndrome de Down (SD) se caracteriza por retraso mental, alteraciones motoras y la aparición de neuropatología tipo Alzheimer a edades tempranas. En el adulto con SD, los cambios neurodegenerativos presentan características diferenciales respecto a las observadas en la EA, pese a que comparte sus signos neuropatológicos. Estas diferencias se deben a que el proceso neurodegenerativo es consecuencia específica de las alteraciones en el neurodesarrollo y la plasticidad neuronal. La hipótesis del efecto de dosis génica en el síndrome de Down (SD) se basa en que el desequilibrio de dosis de genes específicos conduciría a niveles elevados de proteínas concretas con consecuencias fenotípicas. Sin embargo sólo algunos de ellos podrán tener consecuencias observables debido a posibles efectos compensatorios. Proponemos que Dyrk1A es uno de esos genes dosis-sensibles que en base a su patrón de expresión y a los sustratos de fosforilación identificados, podría participar en las alteraciones motoras, cognitivas y el proceso neuropatológico tipo EA en personas con SD a través de la afectación de la neuritogénesis y la neuroplasticidad.DYRK1A es un gen del cromosoma 21 que codifica para una serin-treonin quinasa de actividad dual que se expresa principalmente en médula espinal, cerebelo y núcleos motores relacionados y en estructuras prosencefálicas. Recientemente se ha implicado Dyrk1A en procesos de proliferación y diferenciación neuronal, en la regulación del ciclo celular, en aprendizaje y memoria y en procesos neurodegenerativos.El objetivo general de esta Tesis Doctoral consiste en determinar el papel del gen DYRK1A en el proceso degenerativo presente en el SD, tanto durante el neurodesarrollo como el asociado al envejecimiento y sus mecanismos patogenéticos, utilizando como aproximación experimental modelos de ratón genéticamente modificados con diferente dosis de este gen. Se aborda la descripción y caracterización del impacto de cambios en los niveles de expresión de Dyrk1A en el proceso neurodegenerativo y se estudian los aspectos mecanísticos que posiblemente subyacen a estos procesos, tanto a nivel neuroquímico, centrándonos en dos sistemas de neurotransmisión específicamente relacionados con función cognitiva, como a nivel celular, donde principalmente abordamos aspectos relacionados con neuritogénesis.Nuestros resultados implican a Dyrk1A en el deterioro asociado a la edad de la función motora y la memoria reciente, posiblemente a través de la alteración en los sistemas de neurotransmisión dopaminérgico y colinérgico, respectivamente. Además, sugieren que la sobreexpresión de Dyrk1A tiene consecuencias patogenéticas en los procesos de neuritogénesis, probablemente a través de la modulación de la dinámica del citoesqueleto de actina. Estas alteraciones podrían ser responsables de la falta de efectos cognitivo-conductuales observada en ratones transgénicos en el modelo den neuroplasticidad in vivo (enriquecimiento ambiental)., Aging in Down syndrome (DS) is accompanied by neuropathological features of Alzheimer's disease (AD). In fact, a close relationship exists between both pathologies, so that DS has been proposed as a model to study the predementia stages of AD. The goals of this work were to understand the pathogenic mechanism of AD related to genetic environments associated with DS. To this end we focused on a candidate gene overexpressed in DS, Dyrk1A. Dyrk1A is a serin treonin kinase that has been proposed to participate in neurogenesis, cognitive processes and DS neuropathology. We analyzed the role of Dyrk1A in the altered structural plasticity, neurogenic processes and cognitive function during the AD progression in DS, using murine models with different Dyrk1A dosage. Behavioural, histological and neurochemical studies showed a role of Dyrk1A in AD in DS. Our results involved Dyrk1A in the age-associated decline of motor function and recent memory, possible through an alteration in the dopaminergic and colinergic neurotransmitter systems respectively. Moreover, Dyrk1A overexpression had pathogenetic consecuences in the neuritogenesis process, probably affecting the actin cytoskeleton dynamic regulation. Theses alterations might be responsables of the lack of cognitive/behavioural effects observed in transgenic mice in the in vivo neuroplasticity model.

[spa] El síndrome de Down (SD) se caracteriza por retraso mental, alteraciones motoras y la aparición de neuropatología tipo Alzheimer a edades tempranas. En el adulto con SD, los cambios neurodegenerativos presentan características diferenciales respecto a las observadas en la EA, pese a que comparte sus signos neuropatológicos. Estas diferencias se deben a que el proceso neurodegenerativo es consecuencia específica de las alteraciones en el neurodesarrollo y la plasticidad neuronal. La hipótesis del efecto de dosis génica en el síndrome de Down (SD) se basa en que el desequilibrio de dosis de genes específicos conduciría a niveles elevados de proteínas concretas con consecuencias fenotípicas. Sin embargo sólo algunos de ellos podrán tener consecuencias observables debido a posibles efectos compensatorios. Proponemos que Dyrk1A es uno de esos genes dosis-sensibles que en base a su patrón de expresión y a los sustratos de fosforilación identificados, podría participar en las alteraciones motoras, cognitivas y el proceso neuropatológico tipo EA en personas con SD a través de la afectación de la neuritogénesis y la neuroplasticidad. DYRK1A es un gen del cromosoma 21 que codifica para una serin-treonin quinasa de actividad dual que se expresa principalmente en médula espinal, cerebelo y núcleos motores relacionados y en estructuras prosencefálicas. Recientemente se ha implicado Dyrk1A en procesos de proliferación y diferenciación neuronal, en la regulación del ciclo celular, en aprendizaje y memoria y en procesos neurodegenerativos. El objetivo general de esta Tesis Doctoral consiste en determinar el papel del gen DYRK1A en el proceso degenerativo presente en el SD, tanto durante el neurodesarrollo como el asociado al envejecimiento y sus mecanismos patogenéticos, utilizando como aproximación experimental modelos de ratón genéticamente modificados con diferente dosis de este gen. Se aborda la descripción y caracterización del impacto de cambios en los niveles de expresión de Dyrk1A en el proceso neurodegenerativo y se estudian los aspectos mecanísticos que posiblemente subyacen a estos procesos, tanto a nivel neuroquímico, centrándonos en dos sistemas de neurotransmisión específicamente relacionados con función cognitiva, como a nivel celular, donde principalmente abordamos aspectos relacionados con neuritogénesis. Nuestros resultados implican a Dyrk1A en el deterioro asociado a la edad de la función motora y la memoria reciente, posiblemente a través de la alteración en los sistemas de neurotransmisión dopaminérgico y colinérgico, respectivamente. Además, sugieren que la sobreexpresión de Dyrk1A tiene consecuencias patogenéticas en los procesos de neuritogénesis, probablemente a través de la modulación de la dinámica del citoesqueleto de actina. Estas alteraciones podrían ser responsables de la falta de efectos cognitivo-conductuales observada en ratones transgénicos en el modelo den neuroplasticidad in vivo (enriquecimiento ambiental)., [eng] Aging in Down syndrome (DS) is accompanied by neuropathological features of Alzheimer's disease (AD). In fact, a close relationship exists between both pathologies, so that DS has been proposed as a model to study the predementia stages of AD. The goals of this work were to understand the pathogenic mechanism of AD related to genetic environments associated with DS. To this end we focused on a candidate gene overexpressed in DS, Dyrk1A. Dyrk1A is a serin treonin kinase that has been proposed to participate in neurogenesis, cognitive processes and DS neuropathology. We analyzed the role of Dyrk1A in the altered structural plasticity, neurogenic processes and cognitive function during the AD progression in DS, using murine models with different Dyrk1A dosage. Behavioural, histological and neurochemical studies showed a role of Dyrk1A in AD in DS. Our results involved Dyrk1A in the age-associated decline of motor function and recent memory, possible through an alteration in the dopaminergic and colinergic neurotransmitter systems respectively. Moreover, Dyrk1A overexpression had pathogenetic consecuences in the neuritogenesis process, probably affecting the actin cytoskeleton dynamic regulation. Theses alterations might be responsables of the lack of cognitive/behavioural effects observed in transgenic mice in the in vivo neuroplasticity model.