Altered Reactivity to Threatening Stimuli | Patricia Karkušová
Altered reactivity to threatening stimuli in Drosophila models of Parkinson’s disease, revealed by a trial-based assay
Authors: Patricia Karkušová (1,2), Márton Kajtor (2), Viktor A. Billes (2), Bálint Király (2), Tibor Kovács (2), Hannah Stabb (2), Katalin Sviatkó (2), Andor Vizi (2), Eszter Ujvári (2), Diána Balázsfi (2), Sophie E. Seidenbecher (2), Duda Kvitsiani (2), Tibor Vellai (2)
Supervisor: Balázs Hangya (2)
(1) Biomedical Center, Faculty of Medicine in Pilsen, Charles University (2) Laboratory of Systems Neuroscience, Institute of Experimental Medicine, Budapest, Hungary
State-of-the-Art: To better understand behavioral manifestations of neurodegenerative pathologies, it is useful to employ simple, low-cost, and easy-to-maintain model organisms such as fruit flies (Drosophila melanogaster). Fruit flies exhibit a rich behavioral repertoire in response to threatening stimuli, including freezing (or stopping) and various escape behaviors such as jumping, slow or fast take-off, and running. These behaviors can be effectively analyzed in models of neurodegenerative diseases that affect locomotor activity, such as Parkinson’s disease (PD).
Objective: To assess the effectiveness of a single-animal, trial-based behavioral assay in detecting subtle motor impairments through responses to predator-mimicking visual stimuli in Drosophila model of PD and explore the potential role of dopamine receptors in motor behavior regulation.
Material and Methods: In our study, we used a Drosophila model expressing human mutant alleles of parkin (R275W), α-Synuclein (A53T), and dopamine receptor mutants (Dop1R, Dop1R2, DopEcR). Flies were tested using a behavioral apparatus designed to assess responses to predator-mimicking passing shadows. Natural predator-avoidance behavior served as the motivational context for evaluating their motor impairments.
Results & Discussion: Based on our single-animal, trial-based behavioral assay, we found that parkin flies reacted more slowly to predator-like stimuli compared to controls, while α-Synuclein flies exhibited prolonged stopping durations relative to controls. These findings are consistent with the motor and cognitive deficits observed in humans with PD. Additionally, behavioral similarities between parkin and Dop1R and DopEcR mutant flies were also observed, suggesting a potential connection between dopamine receptors and motor activity.
Conclusion: These results suggest that single-trial behavioral analysis can reveal subtle motor deficits in Drosophila’s predator-avoidance behavior and support the idea that dopamine receptors may have specialized and diverse roles in the regulation of motor function.
Study program: Doctoral study – Physiology and Pathological Physiology | Year of study: 4
ID: 1128
Retinal Remodeling in SCA1 Mice | Olena Yakushko
Structural Remodeling of the Retina in Spinocerebellar Ataxia Type 1 Mouse Model
Authors: Olena Yakushko (1), Jan Cendelín (2), Yaroslav Kolinko (1)
Supervisor: Yaroslav Kolinko (1)
(1) Department of Histology and Embryology, Faculty of Medicine in Pilsen, Charles University
(2) Department of Pathological Physiology, Faculty of Medicine in Pilsen, Charles University
State-of-the-Art: Spinocerebellar ataxia (SCA) type 1 is one of the varieties of polyglutamine diseases,
caused by the expansion of the cytosine-adenine-guanine (CAG) repeat in specific genes. SCA1 is characterized by olivopontocerebellar atrophy and ophthalmic abnormalities affecting the optic nerve and retina. Ophthalmological findings correlate with disease duration. Despite mouse models being commonly used to study neurodegenerative diseases, no histological assessment of the retina has been reported in SCA1 mice. That is why further study is necessary to help clarify the extent of retinal involvement and determine whether retinal changes are comparable between animal models and humans.
Objective: The objective of our study was to compare the volumes of retinal layers and the number of cells in retinal cell layers between mice with spinocerebellar ataxia type 1 and healthy mice at 6 and 10 months of age, using stereological techniques.
Material and Methods: The eyeballs samples were obtained from 6- and 10-month-old mice (8 from SCA1 mice and 8 from healthy mice in each age group). Two series of sections were created from each paraffin block by selecting every 50th section. One series was stained with hematoxylin and eosin to analyze the general morphology of the retina and its cells. The second series was stained using antibodies to rhodopsin [1D4] to evaluate the number of rod nuclei in the outer nuclear layer. The total retina volume, the volumes of different retinal compartments, and the number of cells in the outer and inner nuclear, ganglionic layers were determined using stereological techniques. The Mann–Whitney U test was performed to test for between-group effects.
Results & Discussion: It was found that the total volumes of the outer and inner segments of photoreceptor cells were reduced in SCA1 mice at 6 months compared to healthy mice. No statistically
significant changes were observed in the volumes of different retinal layers between healthy mice and SCA1 mice in both age groups, except the volume of nerve fiber layer, where changes were found in a healthy 10-month-old group. Healthy mice showed a statistically significant decrease in cone numbers with age, whereas SCA1 mice exhibited a trend toward a reduction in cone count from 6 to 10 months. The fraction of cones in both the healthy and SCA1 mice groups reduced at 10 months compared to the corresponding groups at 6 months of age. The ganglionic layer cell count in SCA1 mice decreased by 22% (p<0.01) with age.
Conclusion: The observed changes in SCA1 mice retina were manifested as a reduction in the volumes of photoreceptor inner and outer segments at 6 months, fewer cones in the outer nuclear layer, and a decreased number of cells in the ganglionic layer at 10 months of age.
Funding: This study was supported by EMBO, grant number SLG 5433 and by the Cooperatio
Program, research area MED/DIAG.
Study program: Doctoral study – Anatomy, Histology and Embryology | Year of study: 3
ID: 1111
Diazepam Treatment in Lurcher Mice | Nilpawan Roy Choudhury
DIAZEPAM TREATMENT IN A LURCHER MOUSE MODEL OF CEREBELLAR MOTOR AND COGNITIVE AFFECTIVE SYNDROME.
Author: Nilpawan Roy Choudhury (1)
Supervisor: Jan Cendelín (1,2)
(1) Department of Pathological Physiology, Faculty of Medicine in Pilsen, Charles University
(2) Biomedical Center, Faculty of Medicine in Pilsen, Charles University
State-of-the-Art: Various studies have shown the involvement of the cerebellum in non-motor functions, apart from motor coordination. Lurcher mice are one of the best models for studying cerebellar ataxia and behavioral and cognitive impairments resulting from selective olivocerebellar degeneration. Altered behavior of these mice is hypothesized to be caused in part by stress-induced behavioral disinhibition, i.e., inability to inhibit responses to stimuli and to avoid maladaptive behavior in anxiogenic situations. Such behavioral traits may hypothetically influence the performance of Lurcher mice not only in behavioral but also in cognitive and motor tests.
Objective: The study aimed to investigate whether an anxiolytic compound, Diazepam, could reduce signs of behavioral disinhibition and thereby improve pathological behavioral phenotype in Lurcher mice.
Material and Methods: Lurcher and wild-type mice of the B6CBA strain aged 4-5 months were used for the experiments. Wild-type littermates of Lurcher mice served as healthy controls. Mice were treated with GABA agonist Diazepam in two doses (0.5 mg/kg and 1 mg/kg), which should have anxiolytic but not sedative effects in mice according to the literature. Saline (vehicle) was used as a control. The tests were arranged in a three-week protocol that included open-field test, elevated plus maze test, grip strength measurement, Morris water maze test with a hidden and visible goal tasks, and rotarod test. The examination always started 30 minutes after the injection.
Results & Discussion: Performance of control Lurcher mice was worse in the rotarod and grip strength test than in control wild-type mice. Importantly, Diazepam did not affect grip strength, suggesting that muscle function necessary for performance in other tests has not been affected by this unspecific factor. On the other hand, the higher dose of Diazepam worsened the performance of Lurcher mice, but not wild-type mice, in the rotarod test. Diazepam had no significant effect on exploration in the open field and no effect on entering the anxiogenic open arms of the elevated plus maze. In the Morris water maze, the performance of the mice was not changed by Diazepam except for inconsistent reduction of swimming speed.
Conclusion: We have confirmed that low doses of Diazepam had no significant sedative effect on mice. However, Diazepam did not improve the performance of Lurcher mice in the tests. In the rotarod test, Diazepam even exerted a negative effect, although it did not affect muscle strength.
Funding: This work was supported by GAUK project No. 49724 and Cooperatio (NEUR and MED/DIAG research areas).
Study program: Doctoral study – Physiology and Pathological Physiology | Year of study: 2
ID: 1106
Chronic Implantation of Neuropixels Probe on Rat | Amritesh Suresh
Establishing chronic high-density electrophysiology recordings in freely moving rats using Neurop ixels probes.
Authors: Amritesh Suresh (1,2), Susan Leemburg (1,2)
Supervisor: Karel Jezek (1)
(1) Biomedical Center, Faculty of Medicine in Pilsen, Charles University (2) Department of Pathological
Physiology, Faculty of Medicine in Pilsen, Charles University
State-of-the-Art: The hippocampus plays a key role in spatial navigation and episodic memory
through place cells, which generate cognitive maps via spatially selective firing. CA1 and CA3 contribute differently to memory storage and retrieval. Understanding their interaction requires high-yield recordings from large neuronal populations across regions. Chronic implantation of high-density Neuropixels probes enables stable, large-scale neural data acquisition from multiple hippocampal sub-regions in freely moving rats. This study aims to optimize and validate Neuropixels-based recordings targeting CA1 and CA3 in Long-Evans rats to advance our understanding of hippocampal dynamics during memory retrieval.
Objective: To develop and validate surgical implantation, recording protocols, and analysis
methods for hippocampal Neuropixels recordings in freely moving rats.
Material and Methods: Neuropixels 1.0 probes were implanted in adult Long-Evans rats using
a stereotaxic surgical procedure under anaesthesia. A small craniotomy was performed to access the dorsal hippocampus, and probes were inserted to target both CA1 and CA3 simultaneously.
Post-operative care was provided for 3 days, followed by recovery and habituation in the experimental environment. Neural signals were acquired using Open Ephys acquisition software, alongside video tracking with Bonsai to map location-specific neuronal firing. Both spike and local field potential (LFP) data were analysed. Electrode placement was confirmed using histological analysis following data collection, after perfusion and brain extraction.
Results & Discussion: The Neuropixels implantation protocol was successfully optimized for chronic recordings in freely moving rats. Functional targeting was confirmed by the presence of robust hippocampal theta oscillations and sharp wave ripples in local field potentials. High-quality single-unit activity was reliably captured from both CA1 and CA3 pyramidal layers, with well-isolated waveforms and stable spike clusters across sessions. This approach enables simultaneous monitoring across hippocampal depths, offering enhanced spatial resolution compared to traditional tetrode recordings.
Conclusion: Recordings show stable, low-noise signals, suitable for memory and navigation studies. The optimized method enables investigation of hippocampal circuits and network dynamics, providing a reliable foundation for future experiments examining network dynamics under normal and altered cognitive states.
Funding: Funded by Cooperatio NEUR, and by Grant Agency of The Czech Republic Grant No.
22-16717S.
Study program: Doctoral study – Physiology and Pathological Physiology | Year of study: 2
ID: 1135