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Browsing by study line "Neuroscience and psychobiology"

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  • Fan, Qiuyu (2020)
    Alzheimer’s disease (AD) is a neurodegenerative brain disorder in which the disease process may take decades until the symptoms become evident. To date, no ideal biomarker has emerged that would enable early detection of AD. Environmental and lifestyle factors are thought to affect the risk of developing AD, possibly through epigenetic mechanisms such as DNA methylation (DNAm). DNAm has been shown to differ in the blood and brain of subjects with AD compared with subjects without AD, suggesting that DNAm may be involved in the pathogenic process of AD. This study aims to detect the difference in blood DNAm at baseline between cases who later developed AD and controls who remained AD diagnosis-free during follow-up in a sample selected from a Finnish population-based cohort. Leucocyte genome-wide DNAm was profiled on approximately 850,000 CpG sites by using Infinium MethylationEPIC assay. Each CpG was regressed on the outcome of AD diagnosis during follow-up, controlling for subjects’ age at sampling, sex, smoking status, blood cell counts, working stress level, slide, and array. Specific differentially methylated positions (DMPs) were further explored using pathway analysis. Finally, the methylation level of the candidate gene (APOE) selected from the literature was compared with the sample of this study. After correction for multiple testing, the later diagnosis of AD was not significantly (adjusted p-value < 0.05) associated with methylation level at the baseline at any DNAm site. There was, however, a robust hypomethylation of DMPs among the cases, as 90.9% of the DMPs (p-value < 0.05) were hypomethylated in the case group. The 200 genes annotated by DMPs with the smallest p-values were involved in two neuronal pathways: “Axon guidance associated with semaphorins Homo sapiens” (p-value = 0.0058, adjusted p-value = 0.065) in Panther 2016 and “Semaphorin interactions Homo sapiens” (p-value = 0.00005, adjusted p-value = 0.078) in Reactome 2016. No significant difference existed in DNAm of the candidate gene (APOE) between cases and controls, while cg26190885 at the promoter region of APOE showed nominal significance (p-value = 0.04). In conclusion, no strong evidence was found to support the hypothesis that systemic changes in DNAm are involved in the pathogenesis of AD or that DNAm marks could be detected in blood before the symptoms become evident. A genome-wide pattern of hypomethylation measured by the Infinium MethylationEPIC assay was observed in the case group, serving as a venue for further investigations.
  • Jalkanen, Nelli (2020)
    Mitochondrial aminoacyl tRNA-synthetases (mt-aaRS) catalyse the charging of tRNAs with their cognate amino acids in mitochondria. Mutations in mt-aaRS cause tissue-specific mitochondrial diseases, especially affecting tissues with high energy expenditure like the nervous system, heart, and kidneys. However, disease mechanisms for the heterogeneous group of diseases have not yet been fully elucidated. Harnessing CRISPR-Cas9 genome editing in induced pluripotent stem cells (iPSC) provides an opportunity to model mt-aaRS mutations in vitro and investigate the effects of individual mutations on cellular phenotype. SARS2 encodes mitochondrial seryl tRNA-synthetase, and its c.1347 G>A mutation causes severe childhood-onset progressive spastic paresis. Here, CRISPR-Cas9 ribonucleoprotein (RNP) complex and associated donor template were used to induce homology directed repair (HDR) the genome of iPSC and knock-in the patient mutation. Guide RNAs were designed and tested for efficiency before electroporation into wild type iPSC. Clonal cell lines were made by low-density seeding and manual colony picking. The expression of pluripotency markers was measured by RT-qPCR. RT-qPCR and Western blot measured SARS2 mRNA expression and protein level respectively. The success and precision of genome editing were analysed by Sanger sequencing, comparing the performance of the different guide RNAs, and screening regions of potential off-target genome editing. Two genome-edited iPSC lines with the SARS2 c.1347 G>A mutation were successfully generated to model the patient mutation. The iPSC lines expressed pluripotency markers and contained no off-target genome editing and modelled the patient’s decrease in SARS2 protein level and mRNA expression. More evidence of differentiation ability is needed before differentiation into the affected cell type (motor neurons) and further disease modelling. The efficiency of CRISPR-Cas9 for genome editing, especially harnessing HDR in iPSC, is an area of future research.
  • MacKeith, Ada (2019)
    Sleep difficulties have been on the rise for the past decade. Insomnia and sleep difficulties have associations with an increased risk of overall mortality, as well as with a diverse array of complex diseases, such as coronary heart disease, major depressive disorder, fibromyalgia and Alzheimer’s disease. Epigenomics provides information on how environmental factors influence the genome via epigenetic mechanisms, such as DNA methylation. Thus far, epigenome-wide association studies looking at the effects of sleep disturbances on the methylome have provided evidence of distinctive methylation patterns in insufficient sleep, involving biological processes related to neuroplasticity and neurodegeneration. However, more knowledge is needed to determine how the severity of sleeping difficulties influence the methylome. This thesis investigates the effects of increasing sleep difficulties on DNA methylation with an epigenome-wide association study. The study sample is derived from the Health 2000 general population survey. Subjects were divided into three different groups by their self-reported level of sleeping difficulty, and methylation measurements performed from whole blood samples utilizing the Illumina Infinium MethylationEPIC kit, encompassing >850,000 CpG sites. To identify differentially methylated sites, a multivariable regression model was used with age, gender, smoking, alcohol use, cell type distribution and plate and array data as covariates. None of the differentially methylated CpG sites identified remained significant after multiple testing correction. To gain more information regarding which biological processes the methylated sites may be part of, those CpG sites with an uncorrected p-value of <0.0005 were subjected to pathway analysis. Notable significant pathways included oxytocin- and serotonin receptor-mediated signalling pathways and Alzheimer’s disease-amyloid secretase pathway. Altogether, six pathways remained significant after multiple testing correction, with a total of 12 different genes appearing in them. Furthermore, a post-hoc regression analysis was conducted between these 12 genes and their corresponding CpG sites, and health-related quality of life questionnaire responses. Significant results included associations between sleep, and discomfort and symptoms (including pain). As an additional analysis, a database search was conducted to learn more about the genes’ functionality at the level of phenotype. Results included some variant trait associations to sleep, Alzheimer’s disease and cognitive performance. The associations to Alzheimer’s disease and cognitive performance warrant further research with a similar additive model, perhaps with a larger sample.
  • Ahveninen, Lotta (2022)
    Objectives. Ageing is accompanied by neurobiological changes, such as changes in grey matter (GM) volume and cortical thickness, that mediate a gradual cognitive decline, which can, in turn, be potentially offset by stimulating leisure activities. Choir singing is an especially feasible musical activity with positive effects on physiological, psychological, cognitive, and social functioning in old age. Research investigating the effects of choir singing on the ageing brain is limited. As part of the Brain, Ageing, and Vocal Expression (BRAVE) project, this study aimed to investigate the effect of ageing and choir singing on GM structure. Methods. Using a cross-sectional design and voxel-based morphometry (VBM) and surface-based morphometry (SBM), this study compared GM structure between young (20-39 years; n=35), middle-aged (40-59 years; n=34), and old (60-90 years; n=31) participants and investigated the interaction of age and choir singing on GM structure with amateur choir singer (n=54) and controls (n=46). Results and conclusions. Age had a significant and widespread effect on GM structure, with old participants showing lower GM volume and cortical thickness than young (in bilateral sensorimotor, auditory/language, visual, and limbic areas, midbrain, and cerebellum) and middle-aged (in right visual cortex, thalamus, hippocampus and left auditory cortex) participants. Middle-aged participants also showed lower GM volume and cortical thickness than young participants (in bilateral sensorimotor, language, and visual areas, basal ganglia, cerebellum, and right hippocampus and amygdala). These results corroborate the current understanding of neurobiological ageing. No significant interaction of age and choir singing was found on GM structure, which could be explained by methodological factors. Further research is needed to determine whether choir singing can support brain structure or function across healthy ageing.
  • Järvinen, Elli Katariina (2021)
    Ischemic stroke is a complex disease involving multiple pathophysiological mechanisms. To date, many therapeutic intervention strategies such as anti-inflammatory treatments have been tested, but none of them has been successful. Previous studies have shown that mesencephalic astrocyte-derived neurotrophic factor (MANF) improves stroke recovery and increases the expression of phagocytosis related genes. In this study, the phagocytic and inflammatory effect of monocyte chemoattractant protein 1 (MCP-1), macrophage colony-stimulating factor (M-CSF), complement component 3 (C3), adhesion G protein-coupled receptor E1 (ADGRE1), MER receptor tyrosine kinase (MerTK) and mesencephalic astrocyte-derived neurotrophic factor (MANF) on microglia were studied simultaneously for the first time. The phagocytosis related genes were transiently transfected into a microglial cell line and studied in vitro utilizing phagocytosis assay, fluorescence-activated cell sorting, Western blot and enzyme-linked immunosorbent assay. MCP-1, M-CSF and C3a were shown to enhance microglial phagocytosis without inducing a pro-inflammatory response. In addition, MerTK induces phagocytosis and the synthesis of pro-inflammatory cytokines. In conclusion, the real therapeutic potential of MCP-1, M-CSF, C3a and MerTK in stroke treatment should be further characterized and tested in vivo.
  • Harkki, Juliana Sade Maria (2020)
    Background: Alcohol dependence is a chronic severe substance use disorder that has devastating personal and public health consequences. The efficacy of the current pharmacotherapy options for the treatment of alcohol dependence are modest at best, therefore better alternatives are greatly needed. Lysergic acid diethylamide (LSD) has shown promise in treatment of alcohol dependence in several clinical trials. A sigle high dose of LSD has been suggested to have a treatment effect that last for at least six months, indicating a remarkably better efficacy than the currently available methods. LSD itself has been reported to have a low addiction potential. In mouse models, acute LSD has been demonstrated to reduce ethanol consumption. Yet, the mechanism of action behind these effects has remained largely unknown. LSD is an agonist of serotonin’s 5-HT2A and 5-HT2C receptors which have been shown to modulate the dopaminergic activity of the reward circuitry, a crucial brain area in the initiation of addiction. Intracranial self-stimulation (ICSS) is a procedure for a quantitative assessment of reward behavior in animal models. In ICSS, laboratory rodents self-administer electric stimulation to the dopaminergic pathways of the reward circuitry inducing a reinforcing effect similar to drug reward. Aim: The aim of the current body of work was to use ICSS to assess the acute effects of LSD on reward behavior in C57BL/6JRj mice. This was done to improve the understanding of the mechanism of action of LSD and to evaluate whether the ethanol-consumption-reducing effect of LSD in mice is mediated through the reward mechanism. Methods: Bipolar electrodes targeting the medial forebrain bundle were implanted in the brains of C57BL/6JRj mice in a stereotaxic surgery. The animals were trained to acquire the self-stimulation in the discrete-trial current-intensity procedure. First, the possible dose-dependent acute effects were tested with three different doses of LSD. Next, the acute effect of LSD on amphetamine-induced changes in ISCC were tested. Lastly, a small preliminary test on the effects of LSD on lipopolysaccharide (LPS) -induced changes on ICSS were conducted. Results and conclusions: Acute LSD did not affect reward behavior in ICSS on any of the tested doses. Accordingly, LSD did not affect the facilitation of ICSS induced by acute amphetamine. The results of the LPS experiment were likely to be skewed by the development of tolerance to LPS, therefore the evaluation of the possible effect of LSD was not possible. These findings suggest that the previously reported LSD-induced reduction in ethanol consumption in mice, is not mediated through alteration of the reward mechanism. At the same time, these findings provide further evidence supporting the suggestion that LSD itself does not induce facilitation of the reward circuitry needed for the development of addiction.
  • Laukkanen, Liina (2021)
    This study investigated the in vitro and in vivo effects of direct angiotensin II (ANG) receptor type 2 (AGTR2) agonist Compound 21 (C21). The blockade of ANG receptor type 1 (AGTR1) by AGTR1 antagonists has long been associated with antidepressant and anxiolytic effects. Furthermore, it has been suggested that the therapeutic effects of the AGTR1 antagonists are partially dependent on enhancing the signaling through neuroprotective AGTR2. This suggests that as a specific AGTR2 agonist C21 could be used as a potential therapeutic tool to treat mood disorders that would greatly benefit from new effective treatments. Brain-derived neurotrophic factor (BDNF) is a neurotrophic that binds to tropomyosin receptor kinase B (TRKB). This study aimed to test how C21 affects BDNF:TRKB signaling that has been shown to regulate the therapeutic effects of different antidepressants that act on mood disorders. In vitro effects of C21 on BDNF:TRKB signaling were investigated with ELISA in the cortical cell cultures. Acute AGTR2 stimulation significantly elevated the amount of surface TRKB whereas a prolonged treatment of C21 for three consecutive days induced activation of TRKB. Similarly, combined treatment of C21 and a non-therapeutic treatment of BDNF induced TRKB activation, further linking the AGTR2 stimulation by this compound to the BDNF:TRKB signaling. In vivo effects of C21 on conditioned and unconditioned fear were investigated in mice by using contextual fear conditioning and elevated plus-maze (EPM) respectively. The therapeutic effect of C21 protected mice from conditioned fear but failed to provide similar results for unconditioned fear in the EPM. Interestingly, these stress-protective effects of AGTR2 stimulation were lost in the BDNF-deficient animals. To conclude, AGTR2 stimulation by C21 elevates the amount of surface TRKB that enhances the BDNF:TRKB signaling similar to antidepressants, which further leads to the therapeutic, stress-protective effects. Furthermore, these AGTR2-induced effects were absent without exposure to stress or when BDNF was diminished, indicating that both fear conditioning and BDNF are crucially involved. This study suggests that the AGTR2 is indeed a potential therapeutic target for treating mood disorders, and that in the future C21 could be translated for this use. To achieve this result, the cell types that regulate this effect need to be identified.
  • Sokka, Laura (2021)
    Lactase is a digestive enzyme, and its principal function is to break down lactose, a disaccharide found in milk. The main site for lactase expression is the intestines, however, it is also expressed in other tissues, including the brain. Because the primary substrate, lactose, is not present in the central nervous system, it can be assumed that lactase serves another function besides lactose breakdown outside the digestive system. In C57BL/6NCrl mice, lactase expression is higher in the ventral hippocampus after chronic social defeat stress in comparison to controls. This suggests that lactase expression is to some extent affected by stress. Although lactose metabolism is only necessary for mammals, some other animals – including the zebrafish (Danio rerio) – possess a gene that codes for lactase. Research on the zebrafish lactase gene is scarce, and the expression pattern of its two transcripts, the primary lct-201 and the secondary lct-202, is not known. This study focused on measuring lactase expression in adult wild type zebrafish – both on the gene and on the protein level as enzymatic activity. The effect of stress on lactase expression was also examined by applying two different stress models: netting handling stress as a form of physiological stress, and chronic social defeat as a model for psychosocial stress. Real-time polymerase chain reaction (q-RT-PCR) showed lct-201 expression in all five tissues investigated in this study – the forebrain, the mid-hindbrain, higher intestines, lower intestines, and skeletal muscle, whereas lct-202 was only expressed in the higher and lower intestines. The expression level of lct-201 in the muscle was only fifth of that in the lower intestines. Lactase activity assay on the whole brain and whole intestines displayed enzymatic activity in both tissues, with the activity in the intestines being more than seven-fold compared to the brain. q-RT-PCR on both stressed and control fish whole brain and intestines revealed higher lactase expression in the stressed fish intestines, however, the effect was only seen with a primer pair targeting both transcripts simultaneously, and not for either of them separately. Lactase expression was on average approximately 40 % higher in physiologically and 55 % higher in psychosocially stressed fish in comparison to their respective controls. Neither physiological nor psychosocial stress affected lactase expression in the brain. These findings suggest that the two zebrafish lactase transcripts have distinct expression patterns, which might imply different functional roles for lct-201 and lct-202. Furthermore, these results indicate that lactase is expressed in the zebrafish brain, suggesting that it has a specific function in the central nervous system. Based on the findings in this study, lactase gene expression might be connected to experienced stress – both physiological and psychosocial.