Browsing by master's degree program "Master's Programme in Neuroscience"

Tiivistelmä – Referat – Abstract Genetic variations within the MYO16 gene indicate a common predisposition to severe psychiatric, neurocognitive and neurodevelopmental disorders (NDD), as well as bipolar disorders (BD) and schizophrenia spectrum disorders (SSD). Myosin XVI’s ability to regulate actin and its involvement in cytoskeleton remodeling highlights the protein’s significance in neuronal circuitry development and signaling. Mutations in actin regulator protein-encoding genes like MYO16 have been found to shift cytoskeletal dynamics, as well as cause irregular dendritic spine and excitation/inhibition (E/I) synapse phenotypes. Interestingly, altered actin dynamics and E/I synapse dysregulation are two commonly detected molecular deficits associated with neuropathologies, namely autism spectrum disorders (ASD), SSD, and intellectual disability (ID). Therefore, synaptic E/I profiles are good candidates for investigating the neuropathologies they accompany, and also for revealing potential functional abnormalities. Hence, we determined that quantifying the levels of inhibitory synaptic proteins VGAT and gephyrin is the most suitable approach to investigate inhibitory synapse profiles and their relation to pathologies. Specifically, we investigated how microRNA (miRNA)-mediated myosin XVI protein knockdown (KD) affects pre- and postsynaptic inhibitory synapse density in rat primary hippocampal neurons. We achieved this by analyzing the density of VGAT and gephyrin puncta, signifying pre- and postsynaptic inhibitory synapses, respectively, and also by measuring their diameter to determine differences in inhibitory synapse size. Moreover, we quantified and assessed inhibitory synapse density and size differences between groups by comparing Myo16 KD-plasmid expressing hippocampal neurons to scrambled control cells. Common for both Myo16 KD plasmids was the active suppression of myosin XVI by 33%. However, Myo16 KD plasmids did not affect inhibitory synapse density and size to the same degree. Specifically, there was a significant reduction of inhibitory synapse density in the Myo16 KD3-plasmid expressing neurons, yet, no changes were observed in Myo16 KD5-plasmid expressing neurons. Finally, pre- and postsynaptic inhibitory synapse size differences were not significant between groups for either Myo16 KD plasmid when compared to scrambled control. Aberrant actin cytoskeleton remodeling, as well as altered E/I synapse ratios may lead to hyper/hypo-transmissive neuronal states or cause E/I imbalance, suggesting a complex relationship between actin regulator genes and inhibitory synapses. Our understanding behind their interplay is fairly limited, thus, gaining insight into the mechanisms associated with altered E/I balance remains the primary aim.

Abstract Introduction: Atherosclerotic cardiovascular diseases (ASCVD) cause the biggest burden on our healthcare system and cause most premature deaths. Risk for ASCVD can be lowered by lifestyle choices and medication, as well as several therapeutics such as ethyl eicosapentaenoic acid (E-EPA) supplementation. Here we aimed to investigate the effect of EEPA intervention on known ASCVD risk factors including circulating lipoprotein levels as well as low-density lipoprotein (LDL) aggregation susceptibility, a new independent risk factor for ASCVD. Study design: A study group of 39 healthy men and women participated in a 4-week long dietary supplement trial with 3.9 g/day of E-EPA. A dose of 75 µg/day of vitamin D was included in the E-EPA capsules. Blood samples were drawn before the trial, at weeks 1 and 4 of the intervention and 1 week after the intervention. The study was an open design where participants’ own baseline measurements were used to measure changes. Outcomes: The mean plasma cholesterol concentration was reduced from 3.8 mmol/l to 3.6 mmol/l (p=0.0038 one-way ANOVA) after one week of E-EPA supplementation and remained the same until the end of study period. This change was followed by a change in plasma LDL (p=0.0028 one-way ANOVA) and triglyceride (p=0.0004 one-way ANOVA) concentrations after four week and one week of E-EPA supplementation, respectively. Vitamin D levels increased on average by 18%, showcasing a lower relative response than seen in other vitamin D trials, which can be attributed to high effective baseline concentrations of vitamin D in our study group and the related negative feedback system. LDL aggregation susceptibility did not significantly change in the entire group. However, we discovered that the change in LDL aggregation susceptibility correlated negatively ( = -0.451, p = 0.0039) with the baseline LDL aggregation susceptibility. Thus, LDL aggregation decreased in participants having aggregation-prone LDL at baseline. This finding highlights a possibility that participants with higher LDL aggregation susceptibility may benefit from addition of E-EPA to their diet.

Neurotrophin, Brain-derived neurotrophic factor (BDNF) and its cognate receptor Tropomyosin receptor kinase B (TrkB), have been concomitantly linked with neuronal plasticity as well as antidepressant mechanism of action. Adult hippocampal neurogenesis involves proliferation and survival of new-born neurons and has been related to antidepressant mechanisms and cognitive improvement. Environmental enrichment (EE) enhances adult hippocampal neurogenesis (AHN) and induces anxiolytic-like effects. This study postulates that EE-living conditions could restore the abnormal serotonergic modulation on AHN of our transgenic mice. In this study, a transgenic mouse line wherein TrkB receptor is compromised from serotonergic neurons and AHN found to be impaired was used. To assess the behavioural effects and the changes in learning and memory tasks produced by 10-weeks of EE, a behavioural battery test was performed. Our results suggested anxiolytic-like effects from EE in the transgenic mice. Likewise, cognitive improvements were also observed in both control and transgenic mice promoted by EE. Moreover, hyperactivity observed in transgenic mice in standard conditions could be rescued, and no phenotypical differences were observed between control and transgenic mice subjected to EE. To further study the effects of EE on AHN, cellular proliferation and survival were studied through the incorporation of BrdU. The results indicate that the abnormal serotonergic regulation of AHN was rescued upon EE-living conditions. Moreover, molecular methods used to measure the alteration of gene expression revealed significant upregulation of genes related to neuronal plasticity and epigenetic modifications. Altogether, these results suggest EE promotes the neuronal plasticity, rescues the impaired regulation of AHN and modulates the genetic expression of the transgenic mice. Findings from this study could provide new insights regarding novel targets that could modulate adult brain plasticity.

The insular cortex has been implicated in the neurocircuitry underlying alcohol addiction. The role of the insular cortex and its projections in regulating ethanol intake in AA (Alko-Alcohol) rats has been studied using chemogenetic tools. Chemogenetic activation of the anterior agranular insula (aAI) in AA rats through excitatory DREADDs expressed in the aAI has been found to decrease ethanol consumption. The aAI projects to the central nucleus of the amygdala (CeA), another brain region involved in the development of addiction, particularly in the withdrawal/negative affect stage. In the current study, we sought to further investigate the role of the aAI and the CeA in regulating voluntary ethanol consumption in AA rats. First, we characterized the efferent projections of the aAI in AA rats by chemogenetically activating the aAI with DREADDs and then measuring c-Fos expression in various regions of interest throughout the brain. Next, we investigated the role of the aAI --> CeA projection in ethanol intake by chemogenetically activating or inhibiting the aAI --> CeA projection using the dual viral Cre-dependent DREADD approach. We examined the effects of this manipulation on voluntary ethanol consumption in AA rats in a two-bottle choice paradigm. Finally, we examined the roles of CeA D1Rs (dopamine receptors) and 5-HT2ARs (serotonin receptors) in regulating ethanol intake by examining the effects of pharmacological agonism or antagonism of these receptors on voluntary ethanol consumption in AA rats. Our results from the first experiment reveal significant activation of brain regions including the posterior agranular insula, the mediodorsal nucleus of the thalamus, and the posterior piriform cortex following chemogenetic activation of the aAI. The projections from the aAI to these regions are potentially important in the aAI circuitry in AA rats and are therefore of interest in future studies on the role of aAI circuitry in ethanol intake. In the second experiment, we found no significant effects of aAI --> CeA projection activation or inhibition on ethanol consumption in AA rats, indicating that this projection may not be a key component in regulating ethanol intake in these rats. Finally, we found no significant effects of pharmacological D1R antagonism, 5-HT2AR antagonism, or 5-HT2AR agonism in the CeA on ethanol intake in AA rats, although there was a non-significant trend towards a dose-dependent decrease in ethanol consumption with increasing dose of the D1R antagonist. Our results reveal new neural projections that should be investigated in future research on the role of the aAI in regulating ethanol intake. Studies on the neurobiology underlying alcoholism may reveal new pharmacological or anatomical targets for treatments of alcoholism in humans.

Nuclear receptor subfamily 5 group A member 1 (NR5A1) is a master regulator of both steroidogenesis and gonadal development. Disruptions of NR5A1 can result in differences in sexual development (DSD). With proven interspecies differences in NR5A1 functioning and human material not being available, human stem cells are one of the most achievable, ethical, and accurate models to study the earliest developmental stages of foetal life. However, in currently existing human stem cell-derived gonadal models the expression of NR5A1 has been insufficient without artificial induction due to the lack of knowledge of its distinct biological mechanisms, endogenous ligands, and co-factors. A functional reporter cell line would enable high throughput microscope screening of differentiation protocols with expressed NR5A1. The aim of this thesis was to generate a functional monoclonal human embryonic stem cell (hESC) reporter line for the gene NR5A1 with Alt-R CRISPR-Cas9 ribonucleoprotein (RNP) complex. Firstly, an efficient guide RNA was determined for NR5A1 by T7 assay, and a homology-directed repair (HDR) donor plasmid was designed based on it. Secondly, monoclonal hESC lines were generated with the Alt-R CRISPR-Cas9 RNP complex knock-in method and HDR donor plasmid via electroporation and single-cell sorting. Finally, monoclonal hESC reporter lines were screened with Touchdown PCR and a functionality analysis based on fluorescence and mRNA expression was performed. Two monoclonal hESC reporter lines H9-NR5A1-eGFP cl. 1 and dual-inducible H9-NR5A1-DDdCas9VP192-eGFP cl. 28 were established by using Alt-R CRISPR-Cas9 RNP complex. However, a functional validation performed on H9-NR5A1-DDdCas9VP192-eGFP cl. 28 cells showed the cell line to be non-functional upon NR5A1 upregulation regardless of the expressed eGFP mRNA detected with RT-qPCR.

Hippocampal place fields play a key role in spatial navigation. New place fields are formed during exploratory behavior through long-term potentiation (LTP) and long-term depression (LTD) of synaptic inputs to place cells located in hippocampal CA1. Recently, a novel form synaptic plasticity termed behavioral time scale plasticity (BTSP) has been demonstrated to occur in CA3–CA1 synapses in vitro. BTSP can potentiate synapses that were active several hundred milliseconds before or after a priming event such as a strong and prolonged somatic depolarization. This plasticity rule could be an important complement to well-established spike timing dependent plasticity (STDP) which associates neuronal inputs with outputs at a time scale less than a few tens of milliseconds. The aim of this thesis was to determine whether high frequency antidromic stimulation can act as a priming event that enables BTSP induction in CA1 pyramidal neurons. The underlying assumption was that antidromic stimulation could prime BTSP via action potential backpropagation. High frequency bursting of CA1 neurons in hippocampal slices was achieved with 100 Hz antidromic stimulation of CA1 axons in the alveus. Schaffer collaterals were stimulated 500 ms before or after CA1 burst firing with intensities that were subthreshold for LTP when unpaired. I found that high frequency firing did not enable LTP induction during either of the two experimental protocols, suggesting that neuronal output alone is insufficient for priming BTSP.

The vagus nerve is the longest nerve of the autonomic nervous system. It innervates, among other organs, the stomach, the lungs and the heart, and it reaches several areas of the brain, including the locus coeruleus and the amygdala. The invasive stimulation of this nerve (vagus nerve stimulation, or VNS) is a currently used method for the treatment of refractory epilepsy and pharmaco-resistant depression (Englot et al. 2011; O’Reardon et al., 2006), but the impact that this technique might have on the brain physiology and functions is still under investigation. Various studies (Frangos et al., 2015; Yakunina et al., 2016; Hansen, 2019) have shown that VNS increases noradrenaline production in the brain, a neurotransmitter that is involved in several cognitive processes, such as sleep and mood control. Furthermore, in a study on patients with epilepsy, by Sun et al. in 2017, VNS appeared to have a clear effect on working memory and emotion-attention interaction. Nevertheless, VNS presents all the risks and potential complications that characterize invasive procedures requiring surgery. Therefore, research is now focusing on safer, non-invasive alternatives, such as transcutaneous vagus nerve stimulation (tVNS). This technique allows to stimulate the nerve through its sensory fibres, located in the cymba and tragus of the ear. The scope of the present study was to see whether tVNS would have the same effects on cognitive and affective functions as VNS. The sample for this single blind placebo-controlled study was composed of 30 healthy subjects between 18 and 45 years old. Exclusion criteria included a history of psychiatric, neurological or cardiovascular diseases. All subjects were asked to complete a computer-based task, the Executive Reaction Times-Test. Throughout the test the subjects alternately received an active or a placebo stimulation, and their brain activity was recorded for the whole duration of the test using a 64-channel EEG cap. The Executive-Reaction Times-Test was chosen for this study because it allows to test multiple executive functions simultaneously. The subjects were presented with a series of stimuli on a screen and were asked to react as fast and accurately as possible to “Go” signals, and to refrain from responding when “NoGo” signals appeared. The test started with a triangle pointing either up- or downwards, followed by a brief pause and a traffic light image. The traffic light showed either a red or a green light and included an emotional distractor in the form of a spider or a flower. The red and green lights were alternately used as “Go” or “NoGo” signals, and the rule changed at each test block. In order to complete the task, subjects needed to keep the image of the triangle in their working memory, stay focused on the stimuli and be ready to react or be able to inhibit any responses, thus several main executive functions are being tested: inhibitory control, working memory, attention and emotion-attention interaction. Active stimulation was delivered through clip electrodes that were attached to the tragus of the left ear, whereas placebo stimulation was delivered through clip electrodes that were attached to the left ear lobe. The subjects were not aware of the difference between the two locations. Only the data of 18 subjects was used for the results analysis, because of technical difficulties with the EEG data (some recordings were too noisy, some presented flat channels). The behavioural data was divided into reaction times and errors, which were separately analysed. The EEG data was used to extract the amplitudes of the ERP peaks N2 and P3. The former is a negative peak visible at 200-350ms; the latter is a positive peak visible at 300-500ms. Previous studies have shown the peaks to be associated with response conflict and inhibition (Falkenstein et al., 1999; Donkers et al., 2004; Smith et al., 2013). The behavioural data analysis did not show any significant effect of stimulation on reaction times or error amounts. The ERP analysis, instead, returned interesting results. We observed a main effect of stimulation (p=0.04) in “NoGo” conditions. There was a significant reduction in the N2P3 amplitude and the N2 amplitude in “NoGo” conditions, with active stimulation compared to placebo. These results seem to suggest that with tVNS, fewer cognitive resources are allocated to resolve the inhibitory task, without worsening the subjects’ performance. The lack of significance in the behavioural results might have been due to a ceiling effect, with the Executive Reaction Times-test being too easy for our sample. Overall, the number of errors was too low to conduct a reliable statistical analysis. Nevertheless, the effects we observed on brain physiology would suggest that further research is needed to explore the actual impact of tVNS on cognitive and affective functions.

Kalvojännitteen syntyyn ja sen muutoksiin liittyvät prosessit mielletään usein haastaviksi aiheiksi oppia ja opettaa. Aiheen opetuksesta ja oppimisesta lukiossa ei juurikaan ole tehty tutkimusta, mutta aiemmat tutkimukset yliopisto-opiskelijoilla ovat osoittaneet, että haasteita esiintyy erityisesti lepokalvojännitteen muodostumisen ymmärtämisessä. Aihetta käsitellään suomalaisissa lukiossa pääasiassa ihmisen biologian kontekstissa vapaavalintaisella kurssilla 4 (LOPS 2015) tai 5 (LOPS 2019) ja aihe onkin yksi ihmisen biologian kurssin keskeisistä sisällöistä oppia ja ymmärtää. Kalvojännitteen synty ja sen muutokset on kuvattu suomenkielisissä lukion biologian oppikirjoissa usein yksinkertais-tetusti ja ajoittain virheellisesti. Tämä saattaa johtaa virhekäsitysten syntymiseen, jolloin opiskeltavaa aihetta ei opita riittävällä tasolla. Siksi tämän maisterintutkielman tavoitteena on selvittää, miten solukalvon sähköistä aktiivisuutta opetetaan suomenkielisissä lukioissa ja millaisia virhekäsityksiä opiskelijoilla aiheesta esiintyy. Näiden tulosten pohjal-ta luodaan kehittämistuotos eli opetusmateriaali biologian aineenopettajien käyttöön, jolla voidaan tukea solukalvon sähköisen aktiivisuuden opetusta ja oppimista. Tutkimusmenetelmänä tutkimuksessa käytettiin kehittämistutkimusta, jossa yhdisteltiin teoreettisen ja empiirisen ongelma-analyysin periaatteita. Teoreettisen ongelma-analyysin kautta pyrittiin kartoittamaan lukio-opetuksen kannalta solukalvon sähköiseen toimintaan liittyvät keskeisimmät seikat, joihin monet virhekäsitykset liittyvät, sekä tarkastelemaan niitä eri näkökulmista. Tämä toteutettiin aiemman tutkimuskirjallisuuden avulla. Empiirisessä ongelma-analyysissä tarkasteltiin sekä lukion biologian oppikirjoja (N=3) että kevään 2021 biologian ylioppilaskoevastauksia (N=400) hermosolun aktiopotentiaalin kulkuun liittyen. Molempia aineistoja analysoitiin laadullisen sisällönanalyysin periaatteiden mukaisesti. Teoreettisesta ja empiirisestä ongelma-analyysistä saatujen tulosten perusteella kehitettiin opetusmateriaalin ensimmäinen versio. Opetusmateriaali luetutettiin läpi maisterintutkielman ohjaajilla ja opetusmateriaalia kehitettiin heiltä saatujen kommenttien perusteella. Jatkokehittämisen tuloksena syntyi opetusmateriaalin toinen ja virallinen versio. Tutkimuksen tulokset osoittivat, että solukalvon sähköinen aktiivisuus on haastava aihe sekä opetuksen että oppimi-sen näkökulmasta. Lukion biologian oppikirjoissa esiintyi epätarkkoja kohtia solukalvon sähköiseen aktiivisuuteen liit-tyen ja nämä epätarkkuudet näkyivät opiskelijoiden ylioppilaskokeen vastauksissa yleisinä virhekäsityksinä. Erityisesti esiin nousi virheellinen käsitys natrium-kaliumpumpun ioneja ”palauttavasta” vaikutuksesta aktiopotentiaalin jälkeen. Toinen yleinen virhekäsitys oli, että uusi aktiopotentiaali on mahdollinen vasta kun kalvojännite on hyperpolarisaation jälkeen palautunut lepotilaan. Tulosten perusteella voidaan todeta, että ainakin osa opiskelijoiden virhekäsityksistä on lähtöisin oppikirjoista eikä näitä virhekäsityksiä ole pystytty korjaamaan opettajan toimesta opetustilanteessa. Ylioppi-laskokeessa tehtävään vastanneista opiskelijoista 11,3 % ei osannut vastata tehtävän ensimmäiseen kysymykseen ollenkaan. Vastauksista, joissa opiskelija oli vähintään välttävällä tasolla onnistunut vastaamaan kysymykseen (N=381), 92,4 % sisälsi yhden tai useamman virhekäsityksen. Virhekäsitysten karsimiseksi paras keino on estää niiden syntyminen. Siksi tämän tutkimuksen perusteella voidaan todeta, että lukion uuden opetussuunnitelman biologian oppikirjat tulisi päivittää sellaisiksi, että virheellistä käsitystä ei pääse syntymään. Tämä ei kuitenkaan pelkästään riitä, vaan myös lukion biologian opettajien tulisi luopua vanhoista opetusdioistaan ja varmistaa, että he opettavat aihetta nykytiedon valossa oikein. Tässä apuna toimii tämän tutkimuksen osana kehitetty opetusmateriaali solukalvon sähköisestä aktiivisuudesta hermosolun aktiopotentiaalin kontekstissa. Jotta opetusmateriaalin toimivuutta ja opiskelijoiden käsitteellistä muutosta voidaan arvioida, jatkotutkimukset ai-heesta ovat tarpeellisia.

Evolving societies force universities to transform from the producers of new information sat in their ivory towers towards the role of entrepreneurial universities. The theme of entrepreneurial universities is widely studied internationally, but studies concerning the University of Helsinki (UH) are scarce. The aim of this thesis is to map the current UH bioentrepreneurship ecosystem and the services it provides. The services were mapped and assessed based on how they match the needs of academic bioentrepreneurs. Measures are also suggested on how to develop the ecosystem. This thesis links strongly to the previous literature on entrepreneurial universities and academic entrepreneurship. Entrepreneurial university as a term encompasses an organisation, which strongly supports and encompasses entrepreneurial action in its different functions. Strong technology transfer and commercialization of research via licensing and spinout company formation, is usually linked to entrepreneurial universities. University spinouts are strongly linked to academic entrepreneurship. In spinouts research results and academic tacit knowledge are transformed into enterprises to produce value. The thesis was conducted as a qualitative case study. For the study UH affiliated entities offering entrepreneurship services and bioentrepreneurs originating from within the university were interviewed. The data was analysed with content analysis methods. The results show that UH bioentrepreneurship ecosystem is just in the beginning with multiple useful services but also with some significant flaws. The most significant obstacles preventing the growth of the ecosystem are the university’s negative culture towards entrepreneurship, non-existent communication about the subject and the absence of relevant supportive networks for academic entrepreneurs. Via changing these the critical mass to enable sustainable ecosystem can be achieved.

Sleep is one of the most vital functions of newborns and infants, and it is essential for neuronal network development. Therefore, long-term sleep disturbances have been associated with growth delays and behavioral disorders. Commonly reported infant sleep disturbances, such as night awakenings and difficulties falling asleep, cause distress to parents. Yet, the development of infant sleep in the home environment has not been fully elucidated due to lack of objective measurement parameters. In the current study, we assessed the feasibility of a motion sensor, attached to wearable pants, and ECG textile electrodes to monitor sleep-related respiration and heart rate of newborns and infants. First, we compared signals recorded by the motion sensor’s measurement channels to the standard respiratory piezo effort belt’s signal during daytime EEG recordings. According to our results, the motion sensor’s gyroscope proved to measure respiratory rate most accurately, while the ECG signal transmitted by the sensor was reliable in interpretable sections. We then provided wearable garments and smartphones to families with infants to assess overnight home-use. Our results indicate that different sleep states could likely be identified based on respiration fluctuation visible in the gyroscope’s signals. Moreover, the wearable system was considered practical and easy to use by the parents. Future studies should focus on validating the sensor with clinically approved measures, in order to train the algorithms to automatically identify different sleep-wake states. By doing so, the wearable sensor could provide information on natural infant sleep structure development over long time periods. Additionally, clinical validation of the sensor may result in the development of a companion diagnostic tool for infant cardiorespiratory and movement disorders.