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Browsing by Subject "KCC2"

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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.
  • Loukasmäki, Säde (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.