Institutional creator:

Instytut Biologii Doświadczalnej im. Marcelego Nenckiego PAN

Contributor:

Urban-Ciećko, Joanna : Supervisor ; Jaworski, Tomasz : Supervisor

Publisher:

Instytut Biologii Doświadczalnej im. M. Nenckiego PAN

Place of publishing:

Warszawa

Description:

[3], 104 ages : illustrations ; 30 cm ; Bibliography ; Summary in English

Degree grantor:

Instytut Biologii Doświadczalnej im. Marcelego Nenckiego PAN

Type of object:

Thesis

Abstract:

Glycogen Synthase Kinase-3β (GSK-3β) was discovered for its role in the regulation of glycogen metabolism. GSK-3β is observed in all tissues and it is involved in the regulation of the activity of multiple proteins and metabolic pathways. Studies on mouse models showed that GSK-3β is important during development of central nervous system and in the adult brain. Transgenic mouse model with overexpression of the constitutively active form of GSK 3β[S9A] in the brain is characterized by behavioral changes such as memory deficits and hyperactivity. In adult transgenic mice, structural changes including decreased brain volume and increased thin spines fraction (considered as immature) in granule cells of dentate gyrus (DG) have been observed. Mechanisms underlying abnormal activity of GSK-3β in synaptic function are not fully understood. Here, we analyzed how constitutively active GSK-3β influences morphology of dendritic spines and excitatory synaptic transmission in granular cells of DG in young (3 week-old) transgenic mice. Microscopic analysis showed that increased activity of GSK-3β led to elongation of dendritic spines without changes of spine density. Next, using whole-cell patch-clamp method, we observed increased inter-event intervals of miniature excitatory postsynaptic currents (mEPSCs) while the event amplitude was not changed. Lack of changes in total spine density together with lower frequency of excitatory events suggested lower number of functional synapses. Therefore, in the next step we analyzed the presence of silent synapses. Silent synapses are non-functional (or immature) excitatory synapses, where N-methylo-D-asparate acid (NMDA) receptor is present with lack of functional α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor. We observed an increase in the fraction of silent synapse in GSK-3β[S9A] mice. These results suggest that increased activity of GSK-3β decreases the stability of AMPA receptors in postsynaptic compartment and/or inhibits synapse maturation.The second aim of this study was to identify whether the abnormal activity of GSK-3β can regulate the expression level of microRNAs (miRNAs) in neurons. MiRNAs (small, non-coding RNAs), are key molecules for proper function of central nervous system. Previous studies showed that GSK-3β can regulate the expression level of microRNAs in cancer cells. Such a link has not been reported in neurons. Therefore, we analyzed miRNA expression in hippocampus of GSK-3β[S9A] mice using next generation sequencing (NGS) by Illumina MiniSeq system. Dysregulation of 24 mature and 71 precursor miRNAs in RNA samples was observed. We chose 4 miRNAs for validation by quantitative polymerase chain reaction (PCR). In transgenic mice, miR-221-5p (miR-221*) expression level was significantly downregulated. Next, to define a role of miR-221* in synaptic plasticity we used wild-type primary hippocampal cell culture. Neurons were transfected with miR-221* inhibitor, and imagining for dendritic spine analysis and mEPSCs recordings were performed. Changes of dendritic spine shape and density were not observed. We found an increase in the peak amplitude of mEPSCs, without changes of inter-event intervals after the application of miR-221* inhibitor. Our results indicate that the downregulation of miR-221* enhances excitatory synaptic transmission in hippocampal neurons. Altogether, overactivity of GSK-3β leads to a reduction of functional synapses in hippocampal granular cells of young mice. Moreover, GSK-3β can regulate miRNA expression level in neurons. In GSK-3β[S9A] mice, the expression level of miR-221* is significantly downregulated and the inhibition of miR-221* in primary hippocampal cell culture leads to changes in excitatory synaptic transmission. Structural and electrophysiological changes observed in GSK-3β[S9A] mice might in turn drive aberrant synaptic plasticity

Detailed Resource Type:

PhD Dissertations

Resource Identifier:

oai:rcin.org.pl:237149

Source:

IBD PAN, call no. 20091

Language:

pol

Language of abstract:

eng

Digitizing institution:

Nencki Institute of Experimental Biology of the Polish Academy of Sciences

Original in:

Library of the Nencki Institute of Experimental Biology PAS

Access:

Open