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Jakub Włodarczyk
Laboratory of Cell Biophysics, Nencki Institute of Experimental Biology, PAS, Warsaw, Poland
Stress resilience is the ability of neuronal networks to maintain their function despite the stress exposure. Using a mouse model we investigate stress resilience phenomenon. To assess the resilient and anhedonic behavioral phenotypes developed after the induction of chronic unpredictable stress, we quantitatively characterized the structural and functional plasticity of excitatory synapses in the hippocampus using a combination of proteomic, electrophysiological, and imaging methods. Our results indicate that stress resilience is an active and multifactorial process manifested by structural, functional, and molecular changes in synapses. We reveal that chronic stress influences palmitoylation of synaptic proteins, whose profiles differ between resilient and anhedonic animals. The changes in palmitoylation are predominantly related with the glutamate receptor signaling thus affects synaptic transmission and associated structures of dendritic spines. We show that stress resilience is associated with structural compensatory plasticity of the postsynaptic parts of synapses in CA1 subfield of the hippocampus.
Monika Puchalska, Magdalena Ziółkowska, Ahmad Salamian and Kasia Radwańska
Laboratory of Molecular Basis of Behavior, Nencki Institute of Experimental Biology PAS, Warsaw, Poland
It is widely believed that synaptic transmission in the hippocampus plays a crucial role in formation of contextual memories. Data obtained in our laboratory implies that PSD-95 (postsynaptic density protein 95)-dependent synaptic plasticity in dCA1 area is required for updating of contextual fear memory. To check how inactivation of dCA1 area affects contextual fear memory extinction, we stereotactically injected the adenoviral vectors (AAVs) encoding DREADD receptor hM4 [AAV2.1:hSyn_hM4_mCherry]. Next we trained mice in contextual fear conditioning (CFC). We found that chemogenetic inhibition of dCA1 impaired contextual fear memory extinction and redacted the levels of synaptic protein, PSD-95. To see whether PSD-95 in dCA1 affects the contextual fear memory we stereotactically injected the lentiviral vectors (LVs) encoding short hairpin RNA (shRNA) silencing PSD-95 expression [H1-shRNA_PSD95-Ub_GFP] in the dCA1 area of young adult male and female mice. Next, we trained mice in CFC and performed electrophysiological recordings in the dCA1 area to assess the functionality of synapses with PSD-95 depletion. Moreover, we tested mice in the IntelliCages system to study spatial choice and extinction of appetitive contextual memories in close-to-ecologic conditions. We found that local depletion of PSD-95 in dCA1 decreased synaptic transmission and impaired the contextual fear memory extinction, but not formation or recall. It also impaired ability to find a reward in a complex context at the initial stage of learning, but had no effect on place preference extinction. Together, our data indicates that PSD-95-dependent synaptic transmission in dCA1 is required for updating contextual memories.
Funding: This work is supported by National Science Centre Grant 2020/38/A/NZ4/00483 to K.Radwańska.
Anbarieh Saadat1, Malgorzata Jasinska2, Emilia Białobrodzka3, Ewa Rodziewicz-Flis3, Damian Flis3, Wiesław Ziółkowski3, Elżbieta Pyza1
1Department of Cell Biology and Imaging, Jagiellonian University, Kraków, Poland,
2Department of Histology, Jagiellonian University Medical College, Krakow, Poland,
3Department of Pharmaceutical Pathophysiology, Medical University of Gdańsk
ALS is an incurable, chronic neurodegenerative disease characterized by a selective death of motoneurons in the motor cortex, brainstem, and spinal cord that control voluntary movements of the muscles. The somatosensory cortex is interconnected with other brain areas including the motor cortex. The positive effect of swimming on the progression of ALS disease and the longevity of mice has already been shown. However, the observed effects were related to training before the appearance of the first symptoms of the disease in mice, while the swimming training after the onset of the disease may have a great practical value for ALS patients. In this project, we examined synaptic plasticity in the brains of ALS mice and the imbalance between excitatory/inhibitory synapses in the somatosensory cortex. Mice were divided into three groups: the early stage of ALS, terminal untrained ALS, and terminal swim trained ALS. The number of excitatory and inhibitory synapses in the somatosensory cortex (barrel cortex) was quantified using transmission electron microscopy. The results showed a significant decrease in the number of excitatory synapses between the early stage of ALS and terminal swim trained mice with ALS, indicating positive effects of swimming on the disease.
Funding: NCN OPUS 20 nr UMO-2020/39/B/NZ7/03366 to WZ and EP.
Bogna Badyra1, Matylda Roszkowska1, Karolina Protokowicz1, Dominika Kurpiewska1, Marcin Barański1, Ewa Liszewska2, Jacek Jaworski2, Leszek Kaczmarek1
1Laboratory of Neurobiology, BRAINCITY, Nencki Institute of Experimental Biology of the Polish Academy of Sciences, Warsaw, Poland
2Laboratory of Molecular and Cellular Neurobiology, International Institute of Molecular and Cell Biology, Warsaw, Poland
Brain organoids provide a great tool to decipher human neurodevelopmental diseases, as they can reflect changes occurring even in the prenatal period. Those diseases are often accompanied by aberrant changes in the formation of dendritic spines harboring excitatory synapses. Yet, none of the studies focused on detailed characterization of dendritic spines in organoids. Herein, we present a novel protocol to visualize these phenomena. Human induced pluripotent stem cells were differentiated into cortical spheroids. On subsequent stages of development, organoids were evaluated for proper differentiation. After day 200 live calcium imaging was performed to analyze their maturation and the spontaneous activity of cells. Next, organoids were evaluated for dendritic spines formation using biolistic delivery of lipophilic dye combined with subsequent immunolabeling of pre- and postsynaptic markers. We show that organoids’ maturation can be manifested by the spontaneous activity of neurons with visible synchronization. This maturation is accompanied by changes in the expression of repertoire of synaptic-related proteins. Importantly, we were able to successfully visualize dendritic spines in neurons within organoids. Furthermore, within spines, we observed the colocalization of pre- or postsynaptic proteins. This method enables a more detailed characterization of complex dendritic spine structure and function in both health and disease.
Funding: This work was supported by the Foundation for Polish Science (MAB/2018/10; `Nencki-EMBL Center of Excellence for Neural Plasticity and Brain Disorders: BRAINCITY').
