Neuropsühhiaatriliste endofenotüüpide seos IgLON adhesioonimolekulidega hiire ajus

Väitekirja elektrooniline versioon ei sisalda publikatsioonePsühhiaatriliste häirete multifaktoriaalse patogeneesi mõistmine on suur väljakutse. Neuropsühhiaatriliste häirete modelleerimine loommudelites annab võimaluse uurida, kuidas närviringete düsfunktsionaalsus põhjustab patoloogiliste fenotüüpide avaldumist. Erinevad ülegenoomsed assotsiatsiooniuuringud (GWAS) ning ekspressiooniuuringud on näidanud IgLON perekonda kuuluvate adhesioonimolekulide (Lsamp, Ntm, Opcml, Negr1, IgLON 5) seost inimese neuropsühhiaatriliste häiretega ning käitumiskatsed Lsamp ja Ntm puudulike hiirtega on näidanud IgLON molekulide osalust emotsionaalse ja sotsiaalse käitumise kujunemises. Funktsionaalsed uuringud on näidanud, et IgLON valgud osalevad närviringete kujunemisel ja toimimisel nii arenevas kui ka täiskasvanud ajus. On teada, et tsütoskeleti dünaamilised ümberkorraldused arenevates neuronites on aluseks neuraalsete ringete kujunemisele, kuid IgLON perekonna molekulide roll arenevate neuronite tsütoskeleti reguleerimises on olnud siiani teadmata. Käesoleva töö eesmärk oli selgitada Lsamp ja Ntm vaheliste vastastoimete ja Negr1 mõju aju struktuurile ja funktsioonidele, kasutades vastavate geenide suhtes mutantseid hiiremudeleid. Analüüsisime neuropsühhiaatriliste häiretega seotud morfoloogilisi, anatoomilisi ja käitumuslikke parameetreid Lsamp−/−, Ntm−/−, Lsamp−/−Ntm−/− ja Negr1−/− hiirtes. Mitmetasandiline lähenemine aitab selgitada, kuidas aju struktuursed kõrvalekalded mõjutavad käitumist. Näitasime, et Lsamp ja Ntm mõjutavad varajast neuriitide väljakasvu ja rakkude jagunemist ning apoptoosi teineteisest sõltuvalt ning samasugused vastasmõjud on jälgitavad ka mutanthiirte käitumuslikes reaktsioonides. Leidsime, et Negr1−/− hiirtel on kõrvalekalded neuritogeneesis, neuroanatoomias ja et nende hipokampuses on vähem inhibitoorseid neuroneid, mis võivad olla sellele hiireliinile iseloomuliku puuduliku sotsiaalse ja tunnetusliku käitumise põhjuseks. Lisaks näitavad käesoleva väitekirja tulemused, et IgLON adhesioonimolekulide toime võib olla sõltumatu rakkudevahelisest adhesioonist. Meie uurimistulemused aitavad mõista, kuidas IgLON adhesioonivalgud, mille geenipiirkonnad on olulised riskilookused paljudele psühhiaatrilistele häiretele, reguleerivad närviringete kujunemist, mõjutades neuronite morfoloogiat ja omadusi ning aju anatoomiat. Neid neuronaalseid muutusi, mis seostuvad muutustega käitumises, võib vaadata kui psühhiaatriliste häiretega seotud endofenotüüpe. Oleme näidanud erinevate IgLON puudulikkusega hiiremudelite sobivust psühhiaatriliste häirete modelleerimiseks ning nende mudelite edasine uurimine aitab neuropsühhiaatriliste häirete kujunemist paremini mõista.Understanding the multifactorial pathogenesis of neuropsychiatric disorders is a considerable challenge. Modelling neuropsychiatric disorders in animals provides us a medium to explore the endophenotypes of these disorders to understand how malfunctioning neuronal circuits manifest as pathological phenotypes. Several genome wide association studies (GWAS) and expression studies have linked IgLON superfamily of cell adhesion molecules (Lsamp, Ntm, Opcml, Negr1, IgLON 5) with neuropsychiatric disorders in humans. Analyses of Lsamp and Ntm deficient mice have shown that these genes are involved in patterning of emotional and social behavior. During development, IgLON cell adhesion molecules assist fundamental neuronal communication and the establishment of circuits through morphological changes in the developing neurons, driven by dynamic rearrangements of the cytoskeleton. The role of IgLON molecules in cytoskeletal regulation during development has remained unknown until now. The goal of the present study was to address the effect of interaction between Lsamp and Ntm and the impact of Negr1 on brain structure and function using deletional mouse models. We studied morphological, anatomical and behavioral parameters related to endophenotypes of neuropsychiatric disorders in Lsamp−/−, Ntm−/−, Lsamp−/−Ntm−/− and Negr1−/− mice. This approach allowed us to gain insight into how structural alterations in the brain can influence manifestations at the behavioral level. We showed that Lsamp and Ntm adhesion molecules interact mutually with each other to coordinate early neurite sprouting, proliferation and apoptosis, which manifest at behavior in adult. Our observation on Negr1−/− mice revealed alterations in neuritogenesis and neuroanatomy, and reduced number of inhibitory interneurons in the hippocampus that may underlie the aberrant social and cognitive behavior. Additionally, we propose that the function of IgLON molecules can exhibit through cell autonomous mechanisms during initiation of neurite sprouting independent of cell-adhesion functions. Our findings expand the understanding of how IgLONs, which are candidate genes for a wide spectrum of psychiatric disorders, are involved in the regulation of neuronal circuits at the level of neuronal morphology and neuronal properties, and how they consequently impact the structural anatomy of the brain. These neuronal alterations that manifest as behavioral alterations can be viewed as endophenotypes of neuropsychiatric disorders. We have demonstrated the suitability of IgLON-deficient mice as models for psychiatric disorders. The future investigation of these models enables better understanding of the pathogenesis and treatment of neuropsychiatric disordershttps://www.ester.ee/record=b524318

Promoter-Specific Expression and Genomic Structure of IgLON Family Genes in Mouse

IgLON family is composed of five genes: Lsamp, Ntm, Opcml, Negr1, and Iglon5; encoding for five highly homologous neural adhesion proteins that regulate neurite outgrowth and synapse formation. In the current study we performed in silico analysis revealing that Ntm and Opcml display similar genomic structure as previously reported for Lsamp, characterized by two alternative promotors 1a and 1b. Negr1 and Iglon5 transcripts have uniform 5' region, suggesting single promoter. Iglon5, the recently characterized family member, shares high level of conservation and structural qualities characteristic to IgLON family such as N-terminal signal peptide, three Ig domains, and GPI anchor binding site. By using custom 5'-isoform-specific TaqMan gene-expression assay, we demonstrated heterogeneous expression of IgLON transcripts in different areas of mouse brain and several-fold lower expression in selected tissues outside central nervous system. As an example, the expression of IgLON transcripts in urogenital and reproductive system is in line with repeated reports of urogenital tumors accompanied by mutations in IgLON genes. Considering the high levels of intra-family homology shared by IgLONs, we investigated potential compensatory effects at the level of IgLON isoforms in the brains of mice deficient of one or two family members. We found that the lack of IgLONs is not compensated by a systematic quantitative increase of the other family members. On the contrary, the expression of Ntm 1a transcript and NEGR1 protein was significantly reduced in the frontal cortex of Lsamp-deficient mice suggesting that the expression patterns within IgLON family are balanced coherently. The actions of individual IgLONs, however, can be antagonistic as demonstrated by differential expression of Syp in deletion mutants of IgLONs. In conclusion, we show that the genomic twin-promoter structure has impact on both anatomical distribution and intra-family interactions of IgLON family members. Remarkable variety in the activity levels of 1a and 1b promoters both in the brain and in other tissues, suggests complex functional regulation of IgLONs by alternative signal peptides driven by 1a and 1b promoters.Peer reviewe

Human-Centred Design in the Public Sector: Case Study on Designing a Traffic Congestion Payment System for the City of Munich

This thesis attempts to make a case for using human-centred design for the purpose of innovating within the public sector. First, an in-depth literature survey is conducted to identify the state-of-the-art, current trends and systems in place for using design in the public sector for the purpose of innovation. Capitalizing on the trend of collaboration between public sector organizations and designers, this thesis is done in collaboration with the city of Munich to solve a problem that affects Munich widely today- traffic congestion. The entire human-centred design process from user research, to iterative design and testing, is followed in order to identify user needs and design a product that relieves user pains and also adheres to the constraints of the problem owners. The human-centred methods used in this thesis include interviews, surveys, ideation workshops, landing page experiments, prototyping, and user testing. In order to understand the impact of collaborating with a designer, interviews are conducted with two actors- one from the city of Munich and one from a private sector partner. The main impact is reported to be the ability of human-centred processes to unearth insights in aspects that traditional problem solving had neglected so far. These aspects include a new way of implementing citizen participation and ensuring accountability and transparency about the expenditure of the congestion charge

Neuroprotection through excitability and mTOR required in ALS motoneurons to delay disease and extend survival.

Delaying clinical disease onset would greatly reduce neurodegenerative disease burden, but the mechanisms influencing early preclinical progression are poorly understood. Here, we show that in mouse models of familial motoneuron (MN) disease, SOD1 mutants specifically render vulnerable MNs dependent on endogenous neuroprotection signaling involving excitability and mammalian target of rapamycin (mTOR). The most vulnerable low-excitability FF MNs already exhibited evidence of pathology and endogenous neuroprotection recruitment early postnatally. Enhancing MN excitability promoted MN neuroprotection and reversed misfolded SOD1 (misfSOD1) accumulation and MN pathology, whereas reducing MN excitability augmented misfSOD1 accumulation and accelerated disease. Inhibiting metabotropic cholinergic signaling onto MNs reduced ER stress, but enhanced misfSOD1 accumulation and prevented mTOR activation in alpha-MNs. Modulating excitability and/or alpha-MN mTOR activity had comparable effects on the progression rates of motor dysfunction, denervation, and death. Therefore, excitability and mTOR are key endogenous neuroprotection mechanisms in motoneurons to counteract clinically important disease progression in ALS

Input coding for neuro-electronic hybrid systems

Liquid State Machines have been proposed as a framework to explore the computational properties of neuro-electronic hybrid systems (Maass et al., 2002). Here the neuronal culture implements a recurrent network and is followed by an array of linear discriminants implemented using perceptrons in electronics/software. Thus in this framework, it is desired that the outputs of the neuronal network, corresponding to different inputs, be linearly separable. Previous studies have demonstrated this by either using only a small set of input stimulus patterns to the culture (Hafizovic etal., 2007), large number of input electrodes (Dockendorf et al., 2009) or by using complex schemes to post-process the outputs of the neuronal culture prior to linear discriminance (Ortman et al., 2011). In this study we explore ways to temporally encode inputs into stimulus patterns using a small set of electrodes such that the neuronal culture's output can be directly decoded by simple linear discriminants based on perceptrons. We demonstrate that network can detect the timing and order of firing of inputs on multiple electrodes. Based on this, we demonstrate that the neuronal culture can be used as a kernel to transform inputs which are not linearly separable in a low dimensional space, into outputs in a high dimension where they are linearly separable. Thus simple linear discriminants can now be directly connected to outputs of the neuronal culture and allow for implementation of any function for such a hybrid system. (C) 2014 Published by Elsevier Ireland Ltd

Alternative Promoter Use Governs the Expression of IgLON Cell Adhesion Molecules in Histogenetic Fields of the Embryonic Mouse Brain

The members of the IgLON superfamily of cell adhesion molecules facilitate fundamental cellular communication during brain development, maintain functional brain circuitry, and are associated with several neuropsychiatric disorders such as depression, autism, schizophrenia, and intellectual disabilities. Usage of alternative promoter-specific 1a and 1b mRNA isoforms in Lsamp, Opcml, Ntm, and the single promoter of Negr1 in the mouse and human brain has been previously described. To determine the precise spatiotemporal expression dynamics of Lsamp, Opcml, Ntm isoforms, and Negr1, in the developing brain, we generated isoform-specific RNA probes and carried out in situ hybridization in the developing (embryonic, E10.5, E11.5, 13.5, 17; postnatal, P0) and adult mouse brains. We show that promoter-specific expression of IgLONs is established early during pallial development (at E10.5), where it remains throughout its differentiation through adulthood. In the diencephalon, midbrain, and hindbrain, strong expression patterns are initiated a few days later and begin fading after birth, being only faintly expressed during adulthood. Thus, the expression of specific IgLONs in the developing brain may provide the means for regionally specific functionality as well as for specific regional vulnerabilities. The current study will therefore improve the understanding of how IgLON genes are implicated in the development of neuropsychiatric disorders

Neural cell adhesion molecule Negr1 deficiency in mouse results in structural brain endophenotypes and behavioral deviations related to psychiatric disorders

Neuronal growth regulator 1 (NEGR1) belongs to the immunoglobulin (IgLON) superfamily of cell adhesion molecules involved in cortical layering. Recent functional and genomic studies implicate the role of NEGR1 in a wide spectrum of psychiatric disorders, such as major depression, schizophrenia and autism. Here, we investigated the impact of Negr1 deficiency on brain morphology, neuronal properties and social behavior of mice. In situ hybridization shows Negr1 expression in the brain nuclei which are central modulators of cortical-subcortical connectivity such as the island of Calleja and the reticular nucleus of thalamus. Brain morphological analysis revealed neuroanatomical abnormalities in Negr1−/− mice, including enlargement of ventricles and decrease in the volume of the whole brain, corpus callosum, globus pallidus and hippocampus. Furthermore, decreased number of parvalbumin-positive inhibitory interneurons was evident in Negr1−/− hippocampi. Behaviorally, Negr1−/− mice displayed hyperactivity in social interactions and impairments in social hierarchy. Finally, Negr1 deficiency resulted in disrupted neurite sprouting during neuritogenesis. Our results provide evidence that NEGR1 is required for balancing the ratio of excitatory/inhibitory neurons and proper formation of brain structures, which is prerequisite for adaptive behavioral profiles. Therefore, Negr1−/− mice have a high potential to provide new insights into the neural mechanisms of neuropsychiatric disorders

Neuronal Growth and Behavioral Alterations in Mice Deficient for the Psychiatric Disease-Associated Negr1 Gene

Neuronal growth regulator 1 (NEGR1), a member of the immunoglobulin superfamily cell adhesion molecule subgroup IgLON, has been implicated in neuronal growth and connectivity. In addition, genetic variants in or near the NEGR1 locus have been associated with obesity and more recently with learning difficulties, intellectual disability and psychiatric disorders. However, experimental evidence is lacking to support a possible link between NEGR1, neuronal growth and behavioral abnormalities. Initial expression analysis of NEGR1 mRNA in C57Bl/6 wildtype (WT) mice by in situ hybridization demonstrated marked expression in the entorhinal cortex (EC) and dentate granule cells. In co-cultures of cortical neurons and NSC-34 cells overexpressing NEGR1, neurite growth of cortical neurons was enhanced and distal axons occupied an increased area of cells overexpressing NEGR1. Conversely, in organotypic slice co-cultures, Negr1-knockout (KO) hippocampus was less permissive for axons grown from EC of β-actin-enhanced green fluorescent protein (EGFP) mice compared to WT hippocampus. Neuroanatomical analysis revealed abnormalities of EC axons in the hippocampal dentate gyrus (DG) of Negr1-KO mice including increased numbers of axonal projections to the hilus. Neurotransmitter receptor ligand binding densities, a proxy of functional neurotransmitter receptor abundance, did not show differences in the DG of Negr1-KO mice but altered ligand binding densities to NMDA receptor and muscarinic acetylcholine receptors M1 and M2 were found in CA1 and CA3. Activity behavior, anxiety-like behavior and sensorimotor gating were not different between genotypes. However, Negr1-KO mice exhibited impaired social behavior compared to WT littermates. Moreover, Negr1-KO mice showed reversal learning deficits in the Morris water maze and increased susceptibility to pentylenetetrazol (PTZ)-induced seizures. Thus, our results from neuronal growth assays, neuroanatomical analyses and behavioral assessments provide first evidence that deficiency of the psychiatric disease-associated Negr1 gene may affect neuronal growth and behavior. These findings might be relevant to further evaluate the role of NEGR1 in cognitive and psychiatric disorders