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Browsing by master's degree program "Neurotieteen maisteriohjelma"

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  • Kallo, Henna (2021)
    During the brain development, GABAergic neurons, also referred as interneurons, migrate tangentially from the subpallium to the pallium. After intracortical dispersion, the interneurons start radial migration towards their final location in the cortex. Although the radial migration of interneurons is extensively studied, mechanisms guiding the migration remain relatively unknown. Here we studied how manipulation of cortical activity affects the radial migration and allocation of the cortical GABAergic neurons in the developing mouse brain. For this purpose, we utilized whisker trimming induced sensory deprivation in GAD67-GFP mice at postnatal days 2-5 (P2-P5) followed by cell counting in brain slices derived from P5 and P10-aged mice. In addition, we performed live-imaging of migrating neurons in organotypic cultures derived from P2 SST-TdTomato and 5HT3aR-GFP mice and cultured for 1 day in vitro. These two mouse lines roughly represent early- and late-born subpopulations of the GABAergic neurons. Live-imaging was accompanied by activity manipulations using different drugs and the Designer Receptors Exclusively Activated by Designer Drugs (DREADD) technology. Analysis of the interneurons’ allocation on the barrel cortex after the unilateral sensory deprivation revealed misallocation of GAD67+ neurons on deep cortical layers of the contralateral hemisphere of the ablation group at P5. Analysis of the tracks from the live-imaged migrating interneurons revealed altered saltatory movement behaviour of 5HT3aR+ interneurons when clozapine-N-oxide (CNO) was used to activate the electroporated GFP-GCaMP3-mCherry-hM3Dq neurons located on L2/3 of the cortex. Moreover, we observed reduced motility of migrating interneurons in the organotypic cultures treated with a KCC2 inhibitor that alters GABAA-receptor mediated transmission. Altogether, our results suggest that activity is important in promoting the radial migration of late-born interneurons during the first days of the postnatal development.
  • Nykänen, Roope (2023)
    Diets of wild owls have been studied by pellets, when the nutritional value of owl prey remains unclear. Fatty acids are an essential part of animal nutrition, acting as energy reserve, building blocks of membranes and precursors for signaling molecules. I studied whether Finnish owl species have species-specific characteristics in the fatty acid profiles of adipose tissue due to different diets and feeding habits. I also compared fatty acid profiles between wild and captive owls to elucidate if the diet of captive owls resembles the diet in the wild. Fatty acids from visceral adipose tissue and a liver were transesterified to fatty acid methyl esters, which were identified and quantified by gas-chromatographs linked to either a mass spectrometry or flame ionisation detector. The fatty acid compositions were compared with Principal Component Analysis and statistical significance of the separations between species, origins and tissues were studied by Soft Independent Modelling of Class Analogies. Differences in the relative concentrations of individual fatty acids and their structural category sums were evaluated by the Kruskal-Wallis test followed by the Wilcoxon rank-sum test. Compared to wild owls, captive owl (snowy owl Bubo scandiacus and Eurasian eagle-owl Bubo bubo) adipose tissues had lower relative concentrations of many long-chain polyunsaturated fatty acids (PUFAs), such as arachidonic acid, eicosapentaenoic acid, and docosahexaenoic acid. The captive owls also had lower n-3 PUFA/n-6 PUFA ratio than the wild owls. I suggest that these differences arise from anthropogenic food given to the prey of the captive owls. In the future, the feeding and breeding of captive owls could be improved and potential health hazards prevented by giving the owls a diet rich in long-chain PUFAs, either by giving the prey more versatile diet or adding supplements to the owl’s diet. The adipose tissues of wild Eurasian eagle-owls contained higher total relative concentrations of monounsaturated fatty acids than the species belonging to the Strix genus. On the other hand, long-chain PUFAs were more prominent in the Strix owls. I suggest that these differences in the fatty acid composition are due to the disparities in feeding behaviour: the Eurasian eagle-owls eat carrions and large prey that cannot be swallowed whole, which may lead to consuming more the body outer parts of the prey. The fatty acid composition of the eagle-owls can also be affected by urban prey that have consumed anthropogenic food. In the future, the fatty acid composition of wild prey should be examined to be able to evaluate the prey composition of owls and its effect on the owl fatty acid profile with physiological consequences, the knowledge that could be used in the future decision-making that supports the conservation of owls.
  • Doutel Figueira, Joana Filipa (2022)
    The general question of this research is how beta oscillations are implicated in stopping an ongoing movement. Previous studies regarding movement cancellation have found a significant increase in beta activity in sensorimotor areas, especially in the form of transient increases in beta oscillations, called beta bursts. However, the functional role of beta band activity in stopping is still unclear, mainly because the behavioural tasks used cannot measure the exact timing when the subjects start the stopping process and therefore it is only possible to infer the stopping time. To resolve this, we used head-fixed rats running on a treadmill while performing a Go/NoGo task. In some NoGo trials, the rat starts to run, realizes the mistake and stops before crossing a fixed distance threshold. These are the events being analyzed, called near-mistake events (N=39,366). We found a single beta burst occurring prior to stopping in all five brain regions analyzed (from 44.2±20.1 ms to 55.8±16.0 ms) and positive correlations of beta burst number and power increase with movement speed before stopping. We also found a single alpha burst prior to and during stopping in all brain regions (from 45.9±20.1 ms to 57.1±19.3 ms), supporting previous studies’ findings of alpha band involvement in inhibitory motor actions. Our findings on beta bursts suggest a causality role in stopping an ongoing movement while our results of alpha bursts need to be further analyzed to understand its functional role.
  • Kantonistov, Mikhail (2023)
    Attention deficit hyperactivity disorder (ADHD) is a neuropsychiatric disorder characterized by inattention, hyperactivity and impulsivity. The symptoms appear in childhood and, if left untreated, can continue into adulthood affecting the quality of life. Currently, diagnosing a child's ADHD relies on subjective questionnaires filled out by a parent and an interview. The detection of changes in brain activity especially during everyday activities could bring important information that could help inform the diagnostics of ADHD. The changes in brain activity in persons diagnosed with ADHD during familiar, everyday events have been studied very little. However, ADHD brain imaging studies done during resting state and simple tasks have found changes in large-scale brain networks. These networks can be studied using functional connectivity approach, where the degree of synchronous activity in different brain regions is used to determine the connection strength between these regions. In this thesis, using functional magnetic resonance imaging, the differences in the functional connectivity of the whole brain between children diagnosed with ADHD (n=17) and controls (n=19) were investigated during a virtual reality task that simulates everyday life (EPELI), watching natural-like videos, and resting state. Connectivity matrices were generated with the NiLearn Toolbox program using Seitzman and colleagues' (2018) 300-area parcellation. The connectivity of the whole brain was examined using Network-Based Statistics. During movie watching the ADHD group showed increased connectivity compared to the control group in a network that included several areas of the motor cortex. This may indicate a role for these regions in the hyperactivity symptoms of ADHD. The same network also included the right superior temporal gyrus, which has previously been linked to impulsivity symptoms in individuals diagnosed with ADHD. During the virtual reality task and resting state, no differences in connectivity were observed between the groups. However, differences between the experimental situations were revealed in several networks when the connectivity was compared within the groups. Many of these networks were very extensive and included several subcortical and cerebellar structures in addition to cortical areas. Both the control and ADHD groups showed increased connectivity in the resting state compared to EPELI. This could possibly be due to the differences in the participants' actions during the task performance. To the best of the author's knowledge, this study is the first to examine the functional connectivity of brains diagnosed with ADHD during naturalistic stimuli. The clear differences between the controls and the ADHD group during movie watching are promising for future naturalistic brain studies. Based on the results, network models are effective in studying the functional connectivity of ADHD under different conditions. However, consideration for similarity of activities during the virtual task could have led to the detection of larger differences.
  • Voipio, Mikko Emil Olavi (2020)
    Nitric oxide (NO) is an important signalling molecule in the brain. NO regulates the function of many proteins by e.g. interacting with tyrosine and cysteine residues, thus inducing post-translational modifications. In animal models, inhibition of NO production triggers behavioural effects similarly to those induced by antidepressant drugs. Receptor tropomyosin-related kinase B (TRKB) has been identified as a key player mediating antidepressant drug (AD) induced effects, and it’s a potential target for NO since it displays multiple potential sites for nitration. Preliminary results from our group indicate that TRKB nitration impairs its signalling, and AD uncouple many proteins from TRKB and reorganizes TRKB protein complex. We examined the effect of selective nitric oxide synthase (NOS) inhibitor N⍵-propyl-L-arginine (NPA) in mice submitted to the contextual fear conditioning and found out that inhibiting NO production with NPA has an antidepressant-like effect on mice. We also found out that AD fluoxetine prevents nitration of TRKB receptors in vivo and antidepressant drugs fluoxetine, phenelzine and imipramine disrupt the interactions of TRKB, NOS1 and NOS1 adaptor protein (CAPON) in co-immunoprecipitation assay. To understand the nature of TRKB and NOS1 interaction, we thus examined the protein domains in NOS1 and TRKB using Uniprot database, and we were unable to identify sites that could interact directly. Literature search for NOS1 adapting proteins followed by Uniprot data mining indicated CAPON as a potential candidate to mediate NOS1: TRKB interaction. Our data shows for the first time that antidepressant drugs disrupt TRKB:CAPON:NOS1 interaction, thus protecting TRKB from NOS1-induced nitration. ADs might induce their behavioural effects by preventing NO-induced impair in TRKB signalling
  • Enberg, Emma (2021)
    Kiinnostus käyttää psykedeelejä, kuten lysergihapon dietyyliamidia (LSD) ja psilosybiiniä, erinäisten psykiatristen sairauksien hoidossa ei ole jättänyt huomiotta päihteiden väärinkäyttöä. Tutkimukset ovat osoittaneet alustavia positiivisia vaikutuksia LSD:n käyttämisessä erinäisten addiktioiden, kuten kokaiini-, nikotiini- ja alkoholiriippuvuuksien hoidossa. LSD:n on raportoitu auttaneen joitain alkoholismista kärsiviä pysymään raittiina jopa 6-12 kuukautta yksittäisen LSD annoksen jälkeen. Valitettavasti näitä tuloksia on hankala tulkita, ja vaikutusten taustalla olevat mekanismit tunnetaan huonosti. Tutkimme hiirimallimme avulla, kuinka yksittäinen LSD annos vaikuttaa ahmimiskäyttäytymiseen. Käytimme sukroosiliuosta ahmivaa eläinmallia palkkionottamiskäyttäytymisen mallintamiseen, mikä on yksi addiktioihin liittyvän käyttäytymisen tunnusmerkeistä. Tutkimuksemme tavoitteena oli selvittää vaikuttaako LSD palkkionottamiskäyttäytymiseen, ja siten mahdollisesti aivojen palkkiojärjestelmään. LSD -annostelu (0,05 ja 0,1 mg/kg, i.p.) vähensi akuutisti sukroosiliuoksen ahmimiskäyttäytymistä, mutta vaikutus loppui viikon kuluessa. Tästä havaitusta akuutista vaikutuksesta huolimatta erot ryhmien välillä eivät olleet tilastollisesti merkittäviä. Täten oletettiin, että nettovaikutukset aivojen palkkiojärjestelmään ovat epätodennäköisiä. Kuitenkin pelkän i.p. injektion (10 ml/kg) havaittiin vaikuttavan veden juomiseen. Havaitsimme merkittävän piikin veden juonnissa injektointipäivänä, mikä palautui normaalitasolle jo seuraavaan päivään mennessä. Nämä tulokset johtivat jatkotutkimukseemme, jossa osoitettiin injektion aiheuttavan piikin vedenjuontiin riippumatta siitä, injektoidaanko saliinia vai LSD:tä. Tätä vaikutusta ei enää havaittu, mikäli injektioita annettiin perättäisinä päivinä, mutta jopa yhden tai kahden päivän väli injektioiden välillä riitti palauttamaan injektion aiheuttaman piikin vedenjuontiin. Koska onnistuimme poistamaan vedenjuontiin aiheutuneen vaikutuksen toistetuilla saliini-injektioilla, eikä vaikutus palautunut injektoitaessa LSD:tä, voimme todeta, että vaikutus liittyi injektiotoimenpiteeseen. Keskeisin havaintomme tässä tutkimuksessa oli, ettei LSD:llä ole merkittävää akuuttia vaikutusta sukroosiliuoksen ahmimiskäyttäytymiseen tässä hiirimallissa.
  • Toissalo, Emilia (2022)
    Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by selective loss of upper and lower motor neurons (MN), which causes progressive muscle weakness and paralysis. ALS leads to death typically from 2 to 4 years after diagnosis. It is important to find more effective treatment options for this devastating disease, as the current treatments can prolong the survival by only a few months. Mesencephalic astrocyte-derived neurotrophic factor (MANF) belongs to an evolutionary conserved neurotrophic factor (NTF) family, whose mode of action differs from classical NTFs. MANF is an endoplasmic reticulum (ER) resident protein and is secreted upon ER stress from the ER, and it can protect the cells from ER stress-induced cell death. MANF has shown to be neuroprotective and –restorative in Parkinson’s disease and stroke rodent models. Adeno-associated viral (AAV) vectors can be used to express therapeutic genes in the target tissues for several months, which lessens the need for repetitive dosing. In this master’s thesis project, we aimed to investigate the neuroprotective effects of intrathecally injected AAV1-MANF gene therapy in a SOD1-G93A mouse model of ALS. We used two different MANF genes; full-length MANF and MANF with deleted ER retention signal (MANF-d), to assess the differences between normal and only secreted MANF. Red fluorescent protein (RFP) was used as a control and to further evaluate the transduction and expression of the viral vectors. Intrathecal injections were performed once on 13 weeks old mice, just before the disease onset. Clinical symptom analyses together with a set of behavioral tests were conducted once a week. Mice were sacrificed at the endpoint of the study when they could no longer use their hind limbs for forwarding propulsion. Immunohistochemical staining was performed on spinal cord paraffin samples, where MN count, microglia activation, and RFP expression were evaluated. AAV1-MANF and AAV1-MANF-d treatments improved the motor behavior of the SOD1-G93A mice one-week post- injection. More specifically, a statistically significant difference was seen in the turning times in the static rods test on two different diameter rods compared to control, but there was no difference between MANF groups. In addition, there was a notable difference between AAV1-MANF and the control group on week 16 rotarod scores. There were no statistically significant differences in other tests, survival of the mice, MN counts, or microglia activation between the groups. RFP expression was detected mainly in the ventral areas of the spinal cord with immunohistochemistry. Our results indicate the potential of MANF gene therapy in the treatment of ALS. Furthermore, we showed that intrathecal AAV1-MANF injections were well tolerated.
  • Mäkelä, Alexandra (2023)
    During recent years obesity has been under extra scrutiny due to its globally rising prevalence, multifaceted effects on the human body and common occurrence of comorbidities. It is estimated that one third of the Finnish population over age 40 will be obese by 2028 (THL, 2022). Consequently, development of mitigation strategies has become a high priority in today’s societies leading to a rising need for new treatments. Several studies have shown how pathological adipogenesis has deleterious effect on brain functionality. The neuropathology of obesity could be explained by increased blood-brain barrier (BBB) leakage, oxidative stress, neuroinflammation and glial activation. Pathologically activated astrocytes (astrogliosis) exhibit phenotypical and functional differences compared to healthy astrocytes, typically exhibiting enlarged cell bodies and swollen cytoskeleton. Astrogliosis has been mainly studied in the context of CNS diseases. Recent studies also shed light on the role of astrocytes in the progression of peripheral diseases including cancer metastasis or inflammation (Ma et al., 2023). However, the active astrocytic profile in obesity is relatively underexplored. In this study we report astroglial phenotypic shifts induced by high-fat diet (HFD) feeding and weight loss (WL). No significant change in GFAP expression was seen between mice that were given an HFD for different durations and their corresponding controls. However, we noted a non-significant trend for increased GFAP expression in response to shorter timepoints (5 days of diet change). This suggested an early astrocytic response to diet, which later normalizes over time. We reported healthy morphologies in astrocytes from chow group exhibiting typical simple thin cytoskeleton with long cell protrusions. Astrocytes in HFD-conditions exhibited reactive phenotypes evidenced by swollen cytoskeletal structures and high GFAP immunofluorescence, extensive lipid droplet (LD) accumulation and upregulated metabolic activity. These observations indicated stressful conditions caused by the diet. Astrocytes in WLconditions exhibited varying phenotypes displaying both reactive and healthy characteristics, slight increase of metabolic activity and lipid accumulation. In addition, we reported different immunofluorescence profiles between glial differentiation promoting marker Meteorin and ER stress regulated cytoprotective marker MANF between the experimental groups. These results show that HFD-induced obesity and consecutive weight loss induce a reactive-like phenotypic shift on astrocytes involving both morphological and functional changes.
  • Niemelä, Miska Aleksanteri (2022)
    Master's thesis project includes the backbone assignment of the human activity-regulated cytoskeleton-associated protein C-lobe (hArc, Uniprot ID: Q7LC44), 7-fluoroindole-based tryptophan-labeling method, and comparing that with the 100% double-labeled and 20%(13C) fractionally labeled samples. The project focuses on the effects of 7-fluoroindole-based fluorotryptophan-labeling. hArc C-lobe has only one tryptophan, which makes the analysis easier. Typically fluorotryptophan-labeling is a costly method – fluorotryptophan itself is very expensive and attaching the fluorine to the tryptophan while expressing is expensive and complicated. Fluoroindolebased labeling circles around the problem, as indole and serine are used in procaryotic systems for tryptophan biosynthesis – meaning that fluoroindole, which is cheap, could be used as an alternative for previous methods. Fluoro-labeled tryptophan is used in protein NMR; for example, in binding studies – fluorine-probes are sensitive, and binding of ligand or protein would move these peaks, indicating binding. This project aims to get an insight into the application of this labeling method. The goal is to see if one could utilize one sample with both (1H, 15N, 13C) labeling and 7-fluorotryptophan labeling for binding and structural studies. However, fluorine is very electronegative, affecting surrounding structures and possibly sequentially nearby amino acids. This possible effect will be observed and determined by comparing the 1H15N-chemical shifts between well-established labeling methods and fluoroindolebased labeling. To determine what amino acids in the protein are affected, if they are affected, will be determined by using the backbone assignment results and the results from the sample comparisons.
  • Garnier Artiñano, Tomás (2021)
    Effective population coding is dependent on connectivity, active and passive postsynaptic membrane parameters but how it relates to information transfer and information representation in the brain is still poorly understood. Recently, Brendel et al. (2020) showed how spiking neuronal networks can efficiently represent a noise input signal. This "D_Model” successfully showed that spiking neural networks can recreate input signal representations and how these networks can be resilient to the loss of neurons. However, this model has multiple unphysiological characteristics, such as instantaneous firing and the lack of units related to physical values. The aim of the present study is to build upon the D_Model to add biological accuracy to it and study how information transfer is affected by biophysical parameters. We first modified the D_Model in the MATLAB environment to allow for the simultaneous firing of the neurons. Using our CxSystem2 simulator in a Python environment (Andalibi et al. 2019), we built a network replicating the one used in the D_Model. We quantified the information transfer of Leaky Integrate-and-Fire units that had identical physiological values for both inhibitory and excitatory units (Comrade class) as well as more biologically accurate physiological values (Bacon class). We used various information transfer metrics such as granger causality, transfer entropy, and reconstruction error to quantify the information transfer of the network. We examined the behaviour of the network while altering the values of the capacitance, synaptic delay, equilibrium potential, leak conductance, reset potential, and voltage threshold. Broad parameter searches showed that no single set of biophysical parameters maximised all information transfer metrics, but some ranges fully blocked information transfer by either saturating or stopping neuronal firing. This suggests theoretical boundaries on the possible electrophysiological values neurons can have. From narrow searches within electrophysiological ranges, we conclude that there is no single optimal set of physiological values for information transfer. We hypothesise that different neuronal types may specialise in transferring different aspects of information such as accuracy, efficiency, or to act as frequency filters.
  • Jalanko, Petri (2021)
    Physical fitness has declined during the last decades in adolescents. Furthermore, several studies have found a positive association between physical fitness and brain volume in adolescents, which is noteworthy since the adolescent brain undergoes substantial changes during growth and maturation. However, despite the importance of the cerebellum on adolescents' cognition and coordination, there remains a paucity of evidence on the associations between physical fitness and cerebellum characteristics. Thus, a cross-sectional approach was used to explore the relationship of cardiorespiratory fitness (CRF), power, speed-agility, coordination and overall neuromuscular performance index (NPI) with total gray matter (GM) volume of the cerebellum as well as lobules VI & VIIb, and crus I volume in 40 (22 girls; 18 boys) adolescents. Peak oxygen uptake (V̇O2peak) was measured by the maximal ramp test on a cycle ergometer, lower limb power was determined with standing long jump (SLJ), speed-agility was assessed with the 10 x 5-m shuttle-run test, upper limb coordination was determined with the Box and Block Test (BBT) and NPI was calculated as the sum of SLJ, BBT and shuttle-run z-values. Lean mass (LM) and body fat percentage (BF%) were measured using a bioelectrical impedance analysis. Cerebellum GM volume, lobules VI & VIIb, and crus I volumes were measured by magnetic resonance imaging (MRI). Results demonstrated that V̇O2peak/LM was negatively associated (β = -.045 P= .014) with cerebellum GM volume. No statistically significant associations were found between SLJ, shuttle-run, BBT scores or NPI and cerebellum characteristics in all participants. However, a poorer shuttle-run time was associated (β = -.363 P = .024) with smaller crus I volume in girls and V̇O2peak/LM was negatively associated (β = -.501 P = .031) with lobule VIIb volume in boys. These findings suggest that, in general, CRF and speed agility are associated with cerebellum characteristics in adolescents and there may be sex differences. The results extend our knowledge of the associations between physical fitness and brain volume, but more studies should be conducted to understand the relationship further.
  • Kousa, Ilari (2023)
    This thesis presents a comprehensive exploration of cerebral palsy, acknowledged as the predominant childhood disability. Traditionally viewed through a narrow lens as primarily a motor disorder, recent investigations have broadened this perception significantly. Beyond motor impairments, cerebral palsy manifests an array of comorbidities spanning sensory, emotional, social, and cognitive domains, reshaping our comprehension of its profound impact on individuals' lives. Challenging the static characterization long associated with cerebral palsy, contemporary research has unveiled a compelling dimension - persistent neuroinflammation. Contradicting the notion of a stable condition, these findings suggest potential progressive aspects. The revelation of persistent neuroinflammation prompts a fundamental reconsideration of cerebral palsy's nature. Should its etiological significance be established, it could revolutionise our understanding, suggesting a dynamic condition evolving over time. Conducted through a rigorous search across Pubmed and MEDLINE databases, this thesis stems from an exhaustive exploration of 900 articles. The literature review was conducted in accordance with the PRISM framework. The literature review provides a comprehensive foundation covering the historical context,pathophysiology, and neuropathology of cerebral palsy. Furthermore, it delves deeply into the aforementioned non-classical perspectives, shedding light on the multifaceted nature of this neurological condition. By synthesising classical and contemporary viewpoints, this study endeavours to broaden the discourse surrounding cerebral palsy, fostering a more inclusive and nuanced comprehension of its complexities. This thesis seeks to bridge the gap between traditional views of cerebral palsy as solely a motor disorder and the evolving understanding of its diverse manifestations across various domains. By integrating insights from multiple disciplines and challenging existing paradigms, it aims to contribute to a more holistic framework for conceptualising cerebral palsy. This integrated perspective aims to enhance not only our theoretical understanding but also the practical implications for interventions and support strategies tailored to the multifaceted needs of individuals living with cerebral palsy.
  • Emre, Dusunceli (2022)
    The degree of neurogenesis in the adult hippocampal dentate gyrus (DG) is the center of the discussion in the field of adult neurogenesis. Although there is an on-going controversy, accumulating evidence suggests that the neural stem cells (NSCs) in the adult human DG are very few. The question remains open as to why there are so few NSCs in the adult human DG when compared with the rodent DG. In order to address these questions, it seems necessary to understand the developmental process of the NSCs in the adult human DG. In this thesis, the neural stem and progenitor cells in the fetal human DG are characterized. In addition to these findings, a semi-automatic method for counting and categorizing cells in their expressions of immunochemistry markers is developed.
  • Liiwand, Maj Britt (2022)
    Chronic stress has been linked to the pathogenesis of various disorders, such as generalized anxiety disorder, depression, and post-traumatic stress disorder (PTSD). Stress-induced hyperexcitability of the basolateral amygdala (BLA) has implications in anxiety-like behavior. Promising evidence points to the direction of GluK1 subunit containing kainate receptors (KARs) having a role in the modulation of GABAergic transmission in the lateral amygdala (LA). The aim of the present study was to investigate whether dysfunction of KARs contribute to stress-induced amygdala hyperexcitability and anxiogenesis in mice. Chronic restraint stress (CRS) is an animal model simulating chronic psychological stress. An in situ hybridization experiment was performed to investigate how CRS affects expression levels of GluK1 in the different neuronal populations in the LA. These data show that CRS leads to downregulation of GluK1 expression in the parvalbumin-positive (PV+) interneurons specifically. Patch clamp recordings of spontaneous inhibitory postsynaptic currents showed that CRS did not affect synaptic GABAergic transmission to the principal neurons in the LA. Lastly, conditional knock-out (cKO) mice that have the Grik1 gene knocked out selectively in the PV-expressing interneurons showed no change in anxiety-like behavior after CRS while their wild-type counterparts demonstrated an increase in anxiety-like behavior observable in the elevated plus maze test. Thus, ablation of GluK1 in PV+ interneurons affects the stress-induced anxiogenesis. Due to low number of animals, it cannot be confirmed yet whether the deletion leads to stress resilience or a phenotype where even regular handling is an aversive experience comparable to physical restraint. GluK1 KAR modulation of PV+ interneuron excitability and its susceptibility to stress-related alterations is only a recently discovered phenomenon, and even though this study provides some insight into the underlying mechanism, further research is needed. Systematic characterization of the mechanism could provide a novel tool for understanding and treating stress-related pathological anxiety, possibly helping patients suffering from anxiety disorders resistant to current treatments available.
  • Lindberg, Maiju (2023)
    As the most common mental disorder, anxiety disorders present a major burden to healthcare worldwide and a challenging problem to overcome for the ones suffering from it. Recently, researchers have started to recognize that the relationship between sleep and anxiety disorders is bidirectional; disturbed sleep is a potential risk factor for the progression of anxiety and anxiety can lead to sleep disturbances. However, the neural mechanisms underlying anxiety and sleep problems are still poorly recognized. In this study, we used a chronic sleep fragmentation (SF) paradigm to investigate how disturbed sleep alters anxiety-like behavior in mice and what are the potential underlying neuronal mechanisms. This model was chosen because we wanted to focus on a common form of disturbed sleep in humans rather than total sleep deprivation. We measured anxiety-like behavior in the light-dark box and open field tests right after the 2-week SF period and again after a week of recovery. Additionally, we performed immunohistochemical analysis to study prolonged cell activity (transcription factor ∆FosB), parvalbumin (PV) interneurons and perineuronal net (PNN) structures in the medial prefrontal cortex (mPFC) of the mice. Changes in mPFC activity and related brain areas are associated to anxiety in humans and anxiety-like behavior in rodents alike. Similarly, changes in PV interneurons and PNNs, that regulates PV cell function, are associated to anxiety-like behavior. However, PV interneurons and PNNs have not been previously studied in a setting that combines sleep fragmentation and anxiety-like behavior. We found that chronic SF increases anxiety-like behavior in female mice and that this effect persists at least for a week. Conversely, we did not observe significant increase in anxiety-like behavior in male mice. Both female and male mice showed decrease in ∆FosB in the mPFC suggesting that SF treated mice had lower overall levels of cell activity. Similarly, we found that SF treated mice had decreased PV interneuron intensity in both sexes which could indicate changes in the cell activity. However, the pattern of changes in the IHC results was not identical in males and females. Based on the IHC results, we suggest that SF affects neuronal processes in both sexes but the disparity in them could explain the difference in the behavioral effect. This thesis shows that disturbed sleep can lead to increased anxiety-like behavior in rodent models and recognizes potential targets to study the mechanisms behind the phenomena.
  • Lackman, Madeleine Helena (2022)
    Diabetes mellitus is an incurable disease caused by dysfunctional insulin signaling. The brown adipose tissue (BAT) serves as a hotspot for both lipid and glucose consumption and is thus an attractive target for treating metabolic diseases. Newly surfacing evidence suggest that the endothelial cells (ECs) lining the inner layer of vessels might regulate the morphology and function of adipose tissues. Several studies, including our own, suggest that the vessel density is negatively affected by metabolic diseases. As the BAT is an important organ for systemic lipid and glucose metabolism, and as the effects of metabolic diseases on BAT vessels are not adequately explored, I wanted to investigate how the BAT vasculature changes upon early time points of type 1 (T1D) and 2 (T2D) diabetes in this thesis work. To this end, I used mouse models with chemically induced T1D and genetic T2D and characterized these models with immunohistochemical analyses and immunoassays. To explore the transcriptomic landscapes of ECs and adipose stem cells (ASCs), I analyzed scRNAseq data of BAT stromal vascular fractions (SVF), focusing on changes in gene expression and EC-ASC interactions at a transcriptomic level. Also, by using a publicly available single-cell RNA sequencing (scRNAseq) dataset, I compared BAT SVF gene expression to complement the data resulting from our experiments. The results from this work reveal differential angiogenic responses in the T1D and T2D mouse models and open new avenues of research into how these different pathways are activated and how we can take advantage of these differences to treat diseases. All in all, this work will support the efforts in developing better options for future diabetes prevention, diagnosis, and care.
  • Gustafsson, Michelle (2023)
    Adolescent ill-being has in recent years become a prominent health concern globally. Ill-being during adolescence can have negative consequences for future health and wellbeing, as important patterns of health are formed during this time. This highlights the importance of early identification of risk factors and overarching patterns of mental and physical ill-being and arguments for early intervention during adolescence. The aim of this study was to identify risk factors and co-occurrence of subjective ill-being symptoms in the form of depressive symptoms and subjective health complaints. This study also examined whether the subjective ill-being of students was reflected in cortisol patterns in a naturalistic setting. This since stress has been identified as a key etiological factor in ill-being, through the damaging effect of prolonged exposure to elevated cortisol levels. By applying a novel measure of school atmosphere, the study also aimed to examine the potential protective role of the social atmosphere in school on subjective ill-being and cortisol levels. A total of 329 students from eleven Finnish-Swedish upper secondary schools participated in the cross-sectional study by answering a questionnaire. The salivary cortisol samples were collected from a subsample of the participants, with of a total of 209 participant that met the salivary sampling criteria applied in the study. The methodological framework for the statistical analysis of the study consisted of independent samples t-test, ANOVA, Pearson’s correlation, and multiple linear regression. The results showed a higher prevalence of ill-being in girls and second year students. A significant co-occurrence was found for the subjective ill-being measures of depressive symptom and subjective health concerns. The subjective ill-being was however not reflected in the daily cortisol patterns of students in a naturalistic setting. Furthermore, a positive school atmosphere was significantly negatively associated with subjective ill-being of student in the form of depressive symptoms and subjective health complaints. When controlling for covariates, the subjective meaning of school experienced by the students was identified as a significant protective factor against symptoms of ill-being. These findings identify students in need of additional support and highlights the need of applying an overarching view on student ill-being in future adolescent research. Since no associations was found between daily cortisol patterns and subjective ill-being this study contributes to the understanding of HPA axis in early disorder onset. This study also highlights the importance of subjective meaning in a school context and posits increasing the subjective meaning as a prominent strategy to decrease ill-being among Finnish-Swedish upper secondary school students. Further studies are however needed to assess the causality and to examine these relationships further.
  • Nedeczey-Ruzsák, Petra Dalma (2023)
    Normal sex differentiation depends largely on the healthy development of the bipotential gonad, which is identical in both sexes during early stages of embryonic development. Sex differentiation towards the female phenotype is initiated by the expression of pro-ovarian genes, among which Forkhead Box L2 (FOXL2) is an important regulator. Moreover, FOXL2 was found to be one of the genes most widely implicated in female disorders of sex development (DSD). However, there is a lack of understanding regarding its precise role during ovarian differentiation and development. In order to study the gene during early gonadal development, human embryonic stem cells (hESCs) were used as a model. An inducible FOXL2 activation line was generated in vitro, by applying the CRISPR/Cas9 technique in combination with the tetON and destabilized DHFR systems. The cells were also subjected to gonadal differentiation, based on a previously established protocol. The results showed that the establishment of the activation line was successful, and expression of FOXL2 could only be observed in cells that were treated with trimethoprim and doxycycline. Similar findings were observed in the differentiated activator cells, as again only the induced cells expressed FOXL2. On the other hand, both induced and non-induced differentiated cells showed expression of bipotential gonadal marker genes LHX9, EMX2, GATA4 and WT1. However, in the induced cells a lower relative expression of these markers could be observed. Therefore it seems that relative expression of bipotential gonadal markers was affected by FOXL2 activation. The expression of female gonadal marker genes RSPO1, FSHR, WNT4, AMH and FST was not influenced by FOXL2 activation during gonadal differentiation, as most of the markers showed similar levels of expression in both induced and non-induced cells. Therefore further research needs to be conducted to determine optimal time point of FOXL2 activation during differentiation. Nevertheless, an in vitro model could be generated, which could help in the future to further study the role of FOXL2 in gonadal differentiation, and to better understand pathological mechanisms underlying female DSDs.
  • Rappe, Anna (2021)
    Aging is the progressive accumulation of cellular dysfunction, stress and inflammation. The mitochondrial network plays a central role in the maintenance of cellular homeostasis, with a growing body of evidence assigning dysfunctional regulation of this network as cause or effect of age-related diseases including metabolic disorders, neuropathies, various forms of cancer and neurodegenerative diseases. Neuronal sensitivity to changes in energy supply and metabolic homeostasis make neurons especially susceptible to alterations in the mitochondrial network. Mitophagy, a specified form of autophagy, is the selective degradation and quality control mechanism of mitochondria by engulfment and fusion with acidic endolysosomal compartments of the cell. Mitophagy has been extensively characterised in cultured cells and short-lived model organisms. However, our understanding of physiological mitophagy during mammalian aging is unknown. This study utilizes mito-QC mitophagy reporter mice that enable in vivo detection and monitoring of mitochondrial turnover due to the distinct physicochemical properties of the tandem GFP-mCherry reporter. Using cohort groups of young and aged reporter mice, age-dependent alterations of mitophagy were quantified in the cerebellum and the outer nuclear layer (ONL) of the retina. Specific autophagy and mitophagy markers were used to assess the longitudinal alterations in the mitophagic landscape. Images of fixed brain tissue sections were attained by high-speed spinning disc confocal microscopy for the quantitative and histological analysis. This study characterises the longitudinal alterations of mitophagy in distinct regions of the central nervous system (CNS) of mitophagy reporter mice, demonstrating tissue-specific alterations in mitochondrial turnover throughout physiological time. Åldrande kan definieras som den successiva ackumuleringen av cellulär dysfunktion, stress och inflammation. I upprätthållandet av cellens funktioner och homeostas har det mitokondriella nätverket en central roll. Omfattande forskning visar att åldersrelaterade sjukdomar såsom neuropati, ämnesomsättningssjukdomar, olika cancerformer samt neurodegenerativa sjukdomar föranleds av mitokondriell dysfunktion. Neuroner är beroende av oavbruten energitillförsel och upprätthållen metabolisk homeostas, vilket gör dem speciellt mottagliga för förändringar i det mitokondriella nätverket. Mitofagi är en selektiv form av autofagi som degenererar och kvalitetskontrollerar mitokondrier genom att leverera dem till lysosomer där de bryts ned av hydrolytiska enzymer. Den aktuella kunskapen inom regleringen av och mekanismerna bakom mitofagi baserar sig på gedigen forskning av kortlivade organismer och cellkulturer. Däremot är vår kunskap inom åldrandets inverkan på mitofagi i däggdjur begränsad. I denna studie används musmodellen mito-QC vars rapportörgen består av ett binärt GFP-mCherry-komplex som besitter olika fysikaliska och kemikaliska egenskaper, vilket möjliggör upptäckt och analys av mitofagi in vivo. En kvantitativ jämförelse av mitofagi i unga och åldrande möss genomfördes i vävnadssnitt av cerebellum och av det yttre nukleära lagret av retinan. Specifika autofagi- och mitofagimarkörer användes för att utvärdera de longitudinella förändringarna i mitokondriell degenerering. Bilder för kvantitativ och histologisk analys erhölls med höghastighets spinning-disk-konfokalmikroskop. Denna forskning karaktäriserar de longitudinella förändringarna av mitofagi i definierade regioner av det centrala nervsystemet i musmodellen mito-QC och presenterar vävnadsspecifika förändringar i degenereringen av mitokondrier under åldrandets framskridande.
  • Tienhaara, Samu (2021)
    In visual detection, thresholds for light increments are higher than thresholds for light decrements. This asymmetry has been often ascribed to the differential processing of ON and OFF pathways in the retina, as ON and OFF retinal ganglion cells have been found to respond to increments and decrements, respectively. In this study, the performance of human participants in detecting spatially restricted (diameter 1.17 degrees of visual angle) and unrestricted increments and decrements was measured using a two-interval forced choice task. Background light intensities ranged from darkness through scotopic to low photopic levels. The detection threshold asymmetry found in earlier experiments was replicated with local stimuli. In contrast, however, the asymmetry between increment and decrement detection thresholds disappeared with fullfield stimuli. An ideal observer model was constructed to evaluate the role of two factors, Poisson variations and dark noise, in determining detection thresholds. Based on the model, these factors are insufficient to account for the increment-decrement asymmetry.