Browsing by master's degree program "Master 's Programme in Neuroscience"
Now showing items 41-51 of 51
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(2022)Microglia, the resident immune cells of the central nervous system, react to inflammatory stimuli in the brain in a variety of ways. These include migrating to the site of damage and releasing pro- and anti-inflammatory factors. Previous research indicates that these microglial functions require extensive intracellular calcium signaling. Microglial overactivation can exacerbate neuronal damage, especially in cases of chronic inflammation. The ability to modulate the microglial response to damage would therefore be of great clinical relevance. The endoplasmic reticulum (ER) acts as the cell’s main calcium store and regulates cellular calcium levels primarily through the activity of ryanodine receptors (RYR), inositol-triphosphate receptors (IP3R), and the sarco-endoplasmic reticulum calcium ATPase (SERCA) pump. Calcium depletion from the ER is associated with cellular stress and microglial reactivity and therefore the ER may be an important target for modulating the microglial reactive response. The aim of this study is to show whether ER calcium depletion in a microglial cell line causes changes in protein expression, cellular infiltration, and the release of key pro-inflammatory factors. Drugs that block the pumping of calcium from the cytosol via the SERCA pump, such as thapsigargin, effectively induce a state of calcium depletion in the ER. In the present study, treatment with the SERCA pump inhibitor thapsigargin was found to increase SERCA2 expression in BV2, but not SV40, microglial cell lines. Treatment of microglia with thapsigargin was associated with large increases in the release of pro-inflammatory factors IL-6 and TNF-alpha but had no effect on microglial migration.
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(2023)After decades of lull, the use of psychedelics as therapeutical agents has regained both scientific and public attention. The so-called classical psychedelics are currently studied in the treatment of multiple psychiatric conditions, including addiction. The current understanding implies that psychedelics mediate their subjective effects through serotonergic 5-HT2A receptor binding. The activation of the receptor also leads to increases in brainderived neurotrophic factor (BDNF) expression, a critical part of neuroplasticity mechanisms. At a behavioral level, facilitated neuroplasticity can be observed for example as quicker learning. Because addiction often involves associative learning between the pleasure produced by the drug use and the environmental cues, learning away from these associations could help to prevent relapses and enhance recovery. In this study we aimed to assess LSD’s effect on BDNF levels in amygdalar and cortical regions, and their connection to extinction learning in fear and conditioned place preference paradigms. To evaluate time window for enhanced neuroplasticity, we chose two time points for BDNF level measurement, 24 and 48 h after the LSD injections. In addition, we chose both male and female mice for behavioral experiments to study possible differences between the sexes. We did not observe statistically significant differences between the treatment groups in BDNF levels or behavioral experiments after the single LSD injections. Despite that, this study provides perspectives for improving the experimental setups, as well as helps to evaluate still unanswered questions around the connection between psychedelics and neuroplasticity.
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(2022)Research conducted on neural oscillations have paved the way to unravel the complexities of the brain dynamics underlying behavior and cognition. Neuronal oscillations characterize neuronal activity and processing at all spatial scales from neuronal microcircuits to large-scale brain dynamics and hence link cellular and molecular mechanisms to circuit dynamics underlying behavior. Large-scale oscillations and their inter-areal synchronization can be identified from in vivo electrophysiological data from animal models as well as from human magneto- and electroencephalography (M/EEG) data. Large-scale oscillation dynamics identified from human M/EEG data has been critical for resolving whole-brain oscillation dynamics view but is hindered by the indirectness of the measures. In contrast, rodent in vivo electrophysiology has been conventionally used to resolve oscillation dynamics locally in brain microcircuits. Although these measurements yield critical information of the mechanisms behind local oscillation dynamics, they are difficult to link with whole-brain dynamics view obtained from human M/EEG data. The newly established setup at the Neuroscience Center aims overcome these limitations and allows the measurements directly from the brain of awake head-fixed mice with over 1000 channel measuring simultaneously from both cortical and subcortical structures. This Master’s thesis project objective was to obtain proof-of-concept data to characterize oscillation dynamics during resting-state (RS) from awake behaving mice and to investigate whether these dynamics could be modulated by the manipulating E/I balance. More specifically, the current project aimed to investigate the oscillatory profile of the default-mode network (DMN) activity while manipulating the E/I balance with pharmacological mediums. Electrophysiological data was collected from RS activity from awake mice with two µECoG grids comprising together 512 channels and two laminar Neuropixel probes with each consisting 348 channels. The areas of interest were targeted to capture the DMN activity, covering anterior cingulate cortex (ACC), secondary motor cortex (M2), retrosplenial areas, visual cortical layers, pre- and infralimbic areas, hippocampal areas such as CA1 and dentate gyrus as well as lateral and posterior thalamic areas. The network activity was modulated with pharmacological mediums (sedative, stimulant, control) administered in low acute doses to see their effects on the oscillatory profile. Data from four mice were included into this Master’s thesis work and each mouse was recorded first for 30-minute daily baseline, following a 30-minute pharmacological measurement. This Master’s thesis included the data obtained from the µECoG data to the data analysis focusing on the large-scale cortical activity of the DMN. Power spectral density analysis showed a prominent alpha peak, also seen in humans, across condition with a mild decrease in volume in the stimulant condition. Synchronization was assessed with imaginary part of the phase locking value (iPLV), and the results showed increased synchronization in the stimulant condition and decreased in sedative condition in comparison to the control condition. The amplitude correlation coefficient showed also expected results in both pharmacological conditions, namely higher correlation in stimulant and lower in sedative. This project was able to obtain valuable information of the newly established in vivo electrophysiology setup and the results were in line with our expectations. This promising outcome solidifies the translational potential of the setup and its ability to serve as a translational counterpart in numerous research designs in health and disease.
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(2022)Epileptic patients experience spontaneous recurrent seizures and interictal epileptiform discharges that lead to brain injuries, triggering neuroinflammation and waste product accumulation. Due to the detrimental effect of waste products on brain homeostasis, their removal from the central nervous system is (CNS) is crucial. Meningeal lymphatic vessels (mLVs) located in dura matter contribute to CNS clearance by the drainage of metabolites, waste products, and immune cells from subarachnoid space into cervical lymph nodes. Therefore, because of its role in brain homeostasis, the study of mLVs in different neurological conditions and diseases, including TLE, has gotten increased attention in the last decade. In this study, we sought to understand mLVs role in neuroinflammation and changes in rapid eye movement (REM) sleep stage during epilepsy. For this purpose, we induced mLVs ablation followed by kainic acid (KA) epilepsy model in mice. Shortly, animals were inoculated with AAV-VEGFR3-1-4 to induce mLVs ablation and subsequently challenged with KA to induce status epilepticus. Simultaneously, a control group of animals were injected with a sham AAV and later injection of KA. Afterward, spontaneous EEG activity was registered continuously, and data analysed to compare durations of REM sleep. Also, immunohistochemistry of brain samples was performed to investigate neuroinflammatory changes between experimental groups. Ex-vivo analyses of Iba1 and GFAP expression in brain tissue did not show statistically significant changes in neuroinflammation between experimental groups. However, we observed a trend towards lower expression of inflammatory markers in mLVs ablated animals. The analysis of REM sleep duration shows a progressive reduction of this sleep stage in both groups during the first recording period with a subsequent stabilization during the second one. Our data also indicate that mLVs ablated animals present prolonged REM sleep duration compared to the control group. Although this data contradicts our initial hypothesis it is consistent with the well-established negative correlation between neuroinflammation and REM sleep duration. Future studies should consider a deeper analysis of the glial cell profile for a better understanding of the effect of mLVs dysfunction on epileptic pathology. Moreover, the impact of mLVs ablation on REM sleep duration should be characterized in healthy animals.
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(2023)Beta frequency (15-25 Hz) oscillations in the extracellular field potential recorded by cortical EEG and depth electrodes have been connected to stopping. Especially short increases in beta power, so called beta bursts, occur more frequently close to stopping an ongoing movement or when cancelling a planned action. However, there are discrepancies about the causal role of these beta bursts on stopping. Although some studies indicate causality, in others the bursts occur too late for being causal or their number does not increase prior to stopping. One explanation to the disagreement may lie in the behavioral task commonly used to study the neural correlates of action inhibition, the stop signal task. In this task the movement is cancelled before it starts, and actual stopping is thus hidden from the experimenter. Instead, an estimated stop signal reaction time is mathematically modelled. It is likely that this reaction time varies trial by trial, which causes inaccuracy in the results. We were able to define an exact stopping time using head fixed rats running on a treadmill. This enabled us to align brain activity precisely with stopping. With this task, we showed that the number of transient beta bursts increases just prior to stopping. Moreover, the increase correlates with the velocity. These results indicate that beta bursts are causal to stopping. Beta bursts have been noted to be disturbed in Parkinson’s disease and our results may open new doors for early diagnoses or treatments.
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(2022)Sensory systems display a topographical organization, and in the murine somatosensory system there is oneto-one correspondence between individual whiskers and individual cortical columns called barrels. Functional connectivity in the whisker-to-barrel system is formed prenatally and refined after birth, guided by both spontaneous and whisker-evoked activity. GABAergic connectivity emerges already prenatally and includes transient circuits, but the exact role of GABAergic signalling in early development is elusive. The neuronal, major chloride extruder, potassium-chloride cotransporter (KCC2) is heavily upregulated in the cortex during the first two postnatal weeks resulting in the emergence of hyperpolarizing inhibition. However, in cortical interneurons (INs) KCC2 expression can be detected already at the time of birth. The role of this early interneuronal KCC2 expression is unclear. The aim of this thesis was to study the role of KCC2 in the network activity of cortical INs during the perinatal period. Transgenic mice with conditional inactivation of Kcc2 gene, and expression of the calcium indicator GCaMP6f in GAD2+ neurons (INs) were used to image cortical Ca2+ activity. Transcranial widefield Ca2+ imaging in awake head-fixed mice was performed at the day of birth (P0) and showed that spontaneous, but not evoked, activity was significantly reduced in the knock-out animals. Moreover, immunostaining for the activity-induced transcription factor Egr1 showed that thalamic network activity was significantly decreased in the knock-out and heterozygous animals, suggesting involvement of subcortical areas in the decreased cortical activity. Additional experiments are needed to elucidate the role of other mechanisms contributing to the observed change in activity.
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(2022)Abstract Faculty: Faculty of Biological and Environmental Sciences Degree programme: Master’s programme in Neuroscience Study track: Neuroscience Author: Enija Stoka Title: The Role of Meningeal Lymphatic Vessels in the CNS clearance Level: Master’s thesis Month and year: April 2022 Number of pages: 28 Keywords: meningeal lymphatic vessels (mLVs), brain clearance, glymphatic system, perivascular spaces Supervisor or supervisors: Anaϊs Virenque, Francesco Mattia Noe Where deposited: the Helsinki University Library Additional information: - Abstract: The lymphatic system is a drainage pathway for metabolic waste products, soluble proteins and cerebro-spinal (CSF) as well as interstitial (ISF) fluids. Classically, the lymphatic system has been described all over the body, except the central nervous system (CNS) and the retina. This fact created the question of how the brain is being cleared from harmful solutes. The first system described to being responsible for the clearance of the brain was the glymphatic system, and only recently the existence of lymphatic vessels in the meninges (the meningeal lymphatic vessels, mLVs), has been recognized. However, it is still unknown how these two systems interact in removing solutes from the brain. Here, we analyse if the absence of mLVs affects diffusion and clearance of two tracers with low and high molecular weight (3 kDa and 70 kDa), which have been injected intraparenchymally in wild type (WT) and transgenic (TG) mice lacking functional mLVs. Diffusion of 3 kDa dextran tracer in the surrounding tissue was noticeably increased in WT compared to TG mice, associated with an overall decreased accumulation of the tracer in the parenchyma of the mice lacking mLVs. At the same time, we did not observe a genotype difference in the diffusion or clearance of the 70 kDa dextran tracer. Overall, these results indicate that mLVs dysfunction affects the intraparenchymal diffusion and clearance of low molecular weight molecules.
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(2023)In this master’s thesis project, I studied the association of lipid molecules phosphatidylinositol 4-phosphate (PI4P) and phosphatidylinositol 3-phosphate (PI3P) with autophagy in neurons. One of the aims of the study is to determine the level of basal autophagy in primary hippocampal neurons and to come up with a protocol for autophagosome observation without forcing radical changes in cell culture conditions. Other mammalian cells have extremely weak basal autophagy, but they increase it significantly in response to starvation, for example. However, neurons are extremely sensitive to any changes in their surroundings. They change their morphology, behaviour and biochemical properties, and often they simply do not survive. Therefore, the goal is a protocol for successful autophagy observation with minimal external influence. Despite the debate around basal autophagy in neurons, I observed high levels of basal autophagy in neuronal cells incubated in media without supplements. Also, my observations revealed that the inhibition of the last step of autophagosome processing with Bafilomycin A1, was enough to cause the massive accumulation of large autophagosomes. Results demonstrated that primary hippocampal neurons exhibit high levels of basal autophagy, suggesting that on the contrary to other mammalian cells neurons might not have enough potential to increase autophagy when it is induced pharmacologically or by stressful conditions. This would explain why autophagy induction is often claimed to be ineffective for neuronal cultures. The main goal is to observe and compare PI4P presence on autophagosomes in normal conditions and when autophagosome:lysosome fusion is inhibited with Bafilomycin A1. The side goal is to observe PI3P presence on autophagosomes as well. I transfected primary hippocampal neurons with fluorescent probes for PI4P or PI3P as well as for autophagosome-related protein LC3. Localization data was collected with live-cell imaging on a confocal microscope. As expected, PI3P was not detected on autophagosomes located in soma. It is involved in the initial vesicle biogenesis in distal axons but not in later events taking place closer to the cell body. PI4P showed high degree of colocalization with LC3, indicating PI4P presence on autophagosomes, but only when the fusion was presumably inhibited by Bafilomycin A1. These results suggest that PI4P appears on autophagosomes either as a result of compensatory pathway, where autophagosomes fuse with late endosomes instead of lysosomes; or as a molecule normally involved in autophagosome:lysosome fusion. Literature supports the latter explanation, but it cannot be confirmed without further research. These results give an insight into PI4P role in neuronal autophagy and might be relevant for the future research of autophagy disruption and aggregate accumulation in neuronal diseases as a consequence of abnormal lipid signalling, lipid metabolism and transport.
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(2022)Skeletal muscle is the most abundant tissue in the body, accounting for up to 40-50% of total bodyweight. Regeneration of this tissue is dependent on skeletal muscle stem cells, which are termed satellite cells (SCs) based on their anatomical position between the basal lamina and plasma membrane of muscle fibers. SCs exist under homeostatic conditions in a reversible G0 phase of the cell cycle. Quiescent SCs are recognized by the expression of the paired box 7 (Pax7) transcription factor, in the absence of other myogenic transcription factors such as myoblast determination protein 1 (MyoD) or myogenin (MyoG). Quiescent SCs are metabolically less active with a low oxygen consumption rate. They contain less ATP and have few mitochondria with a low membrane potential in comparison to activated SCs. Activated SCs enter the cell cycle and start to proliferate, undergoing metabolic rewiring to primarily utilize glycolysis for energy production. During early activation, there is an increase in mitochondrial content and ATP production, while the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) increase later during active proliferation. Although similar population dynamics, SCs are a heterogenous population of stem cells, with differences in the expression of notch receptors, stem cell markers, ATP and mitochondrial content, which in turn affect the myogenic potential of the cells. Mitochondria are semi-autonomous, double membrane organelles with various regulation within the cell, such as calcium homeostasis, apoptosis, production of metabolic intermediates, reactive oxygen species (ROS) metabolism, and ATP generation through oxidative phosphorylation (oxphos). Differentiation of various other stem cell types is accompanied by an increase in both mitochondrial content and oxidative phosphorylation, with ultrastructural changes that favour this shift in metabolism. The aim of this thesis was to quantify the ultrastructural changes that occur within SC mitochondria during the early proliferative phase, and to implement a method of Correlative Light and Electron Microscopy (CLEM) for identifying and studying subpopulations of SCs. After isolation and during early activation, SCs contain few mitochondria with a diffuse ultrastructure. Classification of the observed mitochondrial phenotypes revealed heterogeneity both within and between timepoints. During later phases of proliferation, there was an increase in the proportion of mature mitochondria, with an increase in cristae density and a decrease in cristae width. Utilizing genetically modified R26-Snaptag-Omp25 x PAX7CreErt2 mice in which recombination with tamoxifen initiates the expression of mitochondrial outer membrane protein 25 (omp25) bound with a SNAP-tag, allowed for specific and temporal labelling of SC mitochondria by fluorescent SNAP substrates. Performing CLEM on fluorescently labelled SC mitochondria enabled their identification during transmission electron microscopy (TEM). In addition to this, temporal labelling of pre-existing (old) and newly imported (young) omp25 revealed a few cells that contained more old mitochondria, with the cristae density being higher in these. While this indicates a correlation between mitochondrial content and ultrastructure within subpopulations of SCs, further studies are needed to validate these early observations.
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(2021)Study-related burnout is a growing problem among university students. Study-related burnout is defined by exhaustion related to studying, a cynical attitude towards studying and feelings of inadequacy. Burnout can be caused by an overtaxing workload and by a lack of personal, social, or material resources. Stress is a natural part of life, but chronic stress can lead to illness and burnout. Students experience stress in their studies and interventions aiming to reduce stress and prevent burnout are therefore necessary. Psychological flexibility increases wellbeing and prevents burnout and is also related to enhanced learning. Acceptance and Commitment Therapy (ACT) is focused on improving psychological flexibility. Good study skills can also prevent burnout. The aim of this study was to examine university students’ experiences of an online ACT-based intervention that aims to increase psychological flexibility and enhance wellbeing and learning. The aim was also to compare the experiences of two groups of students based on their change in study-related burnout during the course. The participants wrote reflective diaries that were qualitatively analyzed using inductive category development and categories that describe students’ experiences of the intervention were formed. The result of this study shows that students had benefited from the course and described effects on both wellbeing and learning. Students in both study groups experienced effects on wellbeing and learning. The importance of peer-support during studying was discussed, which highlights the need for support from peers who experience similar difficulties and concerns. There were also mentions of negative experiences which the students gave as explanations to why a part of the course had not been effective in enhancing their wellbeing and learning. These categories were more often discussed by students whose risk for burnout had increased during the course. Several categories under the themes of positive experiences of effects on wellbeing and learning were more often discussed by students whose risk for burnout had decreased during the course. The result of this study shows that the ACT-based intervention can be regarded as effective in improving wellbeing and studying, and experiences of the intervention were in general positive. Integration of similar interventions in study programs would be beneficial.
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(2023)At visual threshold, the vision relies on catching incident photons. The ultimate limitation of visual sensitivity arises from the quantal nature of light. At night, the uncertainty of photon arrivals differs fundamentally from daylight conditions, where photon flow can be considered continuous, and sets an absolute physical limitation to visual sensitivity. Visual sensitivity has been postulated to be affected by circadian physiological changes. Here, we have shown, that absolute visual sensitivity is under circadian control in light decrement, or quantal shadow, detection in mice. A behavioural visual task of finding a dark stimulus spot was conducted in a white water maze across several background light intensities leading gradually from clearly visible light to darkness. The percentage of correct choices in the task as a function of light intensity was used to measure visual sensitivity, which was remarkably higher nocturnally. Another parameter affecting visual sensitivity was shown to be the decrement size. Mice were more successful in finding the bigger decrements of the three spatial scales used, as well as succeeding in the task better at night. This finding suggests that visual sensitivity is affected by the absolute number of photons, or more precisely, the absolute number of missing photons in contrast to photons of the background illumination.
Now showing items 41-51 of 51