Stability of Place Fields in Murine Hippocampus | Siddharth Baindur
Investigation of place field stability across two distinct contexts in the dorsal CA1 region of the hippocampus using electrophysiology and micro-endoscopy in freely behaving transgenic mice
Authors: Siddharth Baindur (1,2), Susan Leemburg (1,2), Athira Nataraj (1,2), Amritesh Suresh (1,2),
Karel Blahna (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: Hippocampal neurons encode key dimensions of experience, including
space, time, and contextual information, through distinct activity patterns that underlie cognitive
representations. These dynamics are inherently non-linear and critically influence memory
retrieval, with dysfunction potentially contributing to pathological states such as hallucinations
or delusions. This study seeks to optimize and validate the formation of stable place maps using
a transgenic mouse model expressing calcium-sensitive indicator GCaMP6f, to better understand
the neural dynamics of hippocampal encoding during context-specific spatial memory
activation. This novel method has the benefit of spatial resolution of cells as well as acquiring
data of a larger population of cells in the targeted region of interest.
Objective: To study the kinetics of memory trace activation across large populations of cells in
the dCA1 region of the hippocampus, across two distinct contexts, using one-photon calcium
imaging and multi-unit electrophysiology in freely behaving mice.
Material and Methods: GCaMP6f-expressing C57BL/6 transgenic mice from the GENIE Project
were identified using hot-start PCR and gel electrophoresis of tail-derived DNA. A craniotomy
above dCA1 of the hippocampus was followed by either cortical aspiration and GRIN lens implantation
or multi-unit tetrode implantation, both secured with dental cement. Mice underwent a 6-day training protocol, transitioning from a double to a single maze. During experimentation, neural activity was recorded using the nVista miniscope or Axona system, while locomotion was tracked via overhead camera. Calcium data were processed in IDPS (motion correction, SNR enhancement, PCA-ICA), and electrophysiological signals were clustered using Klusta-Kwik. Neural and positional data were aligned using MATLAB scripts to compute ratemaps.
Results & Discussion: Combined analyses from Ca-imaging and electrophysiology revealed
impaired stability of hippocampal spatial representations across distinct contextual environments.
Ca-imaging, despite capturing large cell populations, showed low consistency of place
fields across trials. These findings suggest limitations in context-specific map formation under
the current experimental conditions. Continuous chronic recording during training may clarify
when and how spatial representations emerge. Enhancing trajectory tracking with tools like
DeepLabCut or AnimalMotionViz could improve spatial resolution. Varying analysis parameters
or using alternate software (e.g., CNMFe) may refine cell detection, while MATLAB scripts require
further optimization for accurate data integration.
Conclusion: Further refinement of behavioral training protocol, analytical methodologies-including
data assimilation, accuracy, signal processing-will be necessary to extract and enhance the spatial encoding in mice in order to further experiment on physiological and pathophysiological conditions like psychosis
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: 4
ID: 1136
Laser Soldering for Atraumatic Tissue Fision | Sima Šarčević
Robotic Laser Tissue Soldering for Atraumatic Soft Tissue Fusion Guided by Fluorescent Nanothermometry
Authors: Sima Šarčević (1), Oscar Cipolato (2), Jachym Rosendorf (1), Alex Anthis (2), Inge Herrmann (2)
Supervisor: Václav Liška (1)
(1) Department of Surgery, Faculty of Medicine in Pilsen, Charles University and University Hospital,
Pilsen (2) Department of Mechanical and Process Engineering, ETH Zurich, Switzerland
State-of-the-Art: Minimally invasive surgery (MIS) has revolutionized patient care by reducing
trauma and recovery time. However, suturing soft tissues in MIS remains challenging due to
limited haptic feedback and the fulcrum effect. Laser tissue soldering offers a promising alternative,
enabling atraumatic, seamless tissue fusion. This study introduces fluorescent nanothermometry-
guided laser soldering, integrating temperature-sensitive nanoparticles into protein-based solders for real-time thermal feedback. The technique ensures precise temperature control (60–80°C) to avoid tissue damage while achieving strong, watertight bonds.
Objective: This study aims to develop a feedback-controlled laser tissue soldering system using
fluorescent nanothermometry for precise, atraumatic soft tissue fusion in minimally invasive
and robotic surgery, overcoming limitations of sutures and adhesives.
Material and Methods: The solder paste comprised albumin, gelatin-methacrylate (GelMA),
TiN nanoparticles (photothermal agents), and BiVO₄:Nd³⁺ nanoparticles (fluorescent nanothermometers).
A 750 nm laser heated the paste, while real-time temperature monitoring was
achieved via fluorescence intensity ratio (FIR) analysis. A PID controller modulated laser power
to maintain optimal temperatures (60–80°C). For thermal imaging, a convolutional neural network
(CNN) upscaled low-resolution data from fiber bundles. The system was tested ex vivo
(porcine liver, intestine) and in vivo (porcine model) using robotic and laparoscopic setups. Mechanical
testing compared soldered and sutured liver samples for tensile strength.
Results & Discussion: The feedback-controlled system achieved precise temperature regulation
(<10 s response, no overshooting) and robust tissue bonding. Automated soldering outperformed
manual methods, producing uniform seams without thermal damage. CNN-based thermal imaging accurately reconstructed temperature distributions (ΔTmax = 2.6 ± 1.0°C). In vivo experiments demonstrated seamless integration with laparoscopic tools, with histology confirming strong adhesion and no tissue damage. Tensile strength tests showed soldered liver outperformed sutures due to uniform stress distribution. The technology’s adaptability to robotic systems (e.g., KUKA, Dexter robots) highlights its potential for clinical translation, particularly in fragile tissues like intestines or blood vessels.
Conclusion: Fluorescent nanothermometry-guided laser soldering enables precise, atraumatic tissue fusion in MIS. With automated feedback and robotic compatibility, this method surpasses sutures in bond strength and safety, offering a transformative approach for minimally invasive wound closure.
Study program: Doctoral study – Surgery | Year of study: 3
ID: 1130
Prognostic factors in surgical treatment of HGG | Richard Koleják
Prognostic factors in surgical treatment of high grade gliomas
Authors: Richard Koleják (1), Mraček J. (1), Houfková K. (2), Polívka J. (3), Vítovec M. (4), Švajdler M. (5)
Supervisor: Přibáň V. (1)
(1) Department of Neurosurgery, Faculty of Medicine in Pilsen, Charles University and University Hospital, Pilsen (2) Department of Biology, Faculty of Medicine in Pilsen, Charles University (3) Department of Histology and Embryology, Faculty of Medicine in Pilsen, Charles University (4) Department of Radiology, Faculty of Medicine in Pilsen, Charles University and University Hospital, Pilsen (5) Department of Pathology, Faculty of Medicine in Pilsen, Charles University and University Hospital, Pilsen
State-of-the-Art: High grade gliomas are incurable tumors with fatal prognosis. Number of clinical prognostic factors have been known for many years. The focus has shifted to molecular biomarkers in recent years, which is reflected in the new WHO tumor classification 2021. New molecular methods, such as NGS offer the option to create a personalized molecular tumor profile. Liquid biopsy from blood and cerebrospinal fluid seems to be another promising method, which could be used as one of the modalities for monitoring the course of the disease in the future. In our study we present two groups of patients.
Objective: 1. A retrospective analysis of a group of patients and evaluation of factors impacting the surgical treatment. 2. A prospective analysis group of patients and evaluation of prognostic factors, including methods such as NGS, liquid biopsy and volume measurment.
Material and Methods: The retrospective group consists of 143 patients operated in years 2015-2021. 68 of these patients were lost to followup and thus were not included in the final analysis, which was discussed last year. The prospective group consists of 40 patients operated in years 2022-2024. The analysis evaluates prognostic factors and the effect of these factors on the outcome. Pre- and postoperative volume measurement was also performed. Liquid biopsy from blood and cerebrospinal fluid and evaluation of specified miRNAs is currently in progress.
Results & Discussion: The tumor volume ranged from 1,28ml to 83,08ml, averaging 29,3ml. A radical ressection (>90 % tumoral volume) was achieved in 88,9 % of cases, with an average volume ressection rate of 96,7 % (57,5 – 100 %). Average OS was 364 days. PFS was 210 days. Radical resection (> 90 %) was coupled with higher OS compared to non-radical resection (412 vs 277 days). Average OS with MGMT methylated tumor was higher (449 days), as well as OS following chemotherapy of MGMT met. tumor (701 days). OS was similar in EGFR amplified tumors (366 days). TERT mutated tumors presented with lower OS (311 days). Chromosome 7 and 10 aneuploidy was coupled with lower OS (294 days). Lastly, patients presenting with more than 1 negative molecular marker had an average OS of 264 days.
Conclusion: The surgical results and demographics are consistent with literature. The presence of negative molecular markers is coupled with a lower overall survival times. The last part, liquid biopsy of specified miRNAs encountered in blood and cerebrospinal fluid is currently in progress.
Study program: Doctoral study – Surgery | Year of study: 4
ID: 1129
Bile Duct Reconstruction by Decellularized Tissue | Jan Ševčík
Decellularized bile duct and its partial recellularization by mesenchymal stem cells in vitro – is it suitable for biliary tract surgery?
Authors: Jan Ševčík (1), Maria Stefania Massaro (2), Sima Sarcevic (1), Dan Lehner (2), Martin Faško
(2), Hanna Nochevkina (2), Eliška Zaengerová (2), Anna Štrobachová (2), Ekaterina Panova (2)
Supervisor: Václav Liška (1)
(1) Department of Surgery, Faculty of Medicine in Pilsen, Charles University and University Hospital,
Pilsen (2) Biomedical Center, Faculty of Medicine in Pilsen, Charles University
State-of-the-Art: A bile duct reconstruction after injury or resection is a staple of hepatopancreatobiliary
surgery. Research to find suitable bile duct substitute for anatomical reconstruction
has been conducted over the last 100 years without an optimal result. New methods and
discoveries in Tissue Engineering field offers potential answers to this issue, with potential aplication
for grafts made from decellularized tissue with or without cellular reseeding modification.
Objective: The goal of this study was to verify the possibility of decellularization of allogeneic
porcine extrahepatic bile ducts (eBD), their recellularization by human mesenchymal stem cells
(hMSC), and their usage for common bile duct reconstruction in experimental settings.
Material and Methods: Porcine eBDs were harvested and cryopreserved at -80°. Bile ducts
were decellularized by perfusion with detergents in 4 1-hour cycles. Scaffolds were evaluated for the presence of residual nuclei. For 24 hours, static seeding with hMSC was done in the incubator,
followed by dynamic perfusion seeding in an in-house-developed bioreactor. Live-dead assay and immunofluorescence staining were used to evaluate the presence of living cells on the decellularized bile duct scaffold. An experimental group of six domestic pigs underwent a common bile duct resection with reconstruction by decellularized allogeneic eBD, surviving for 4 weeks to expose complications and show biointegration in the recipient tissues, compared to a control group with common bile duct reconstructed by direct anastomosis.
Results & Discussion: The decellularization process yielded great results in the complete decellularization
of eBD without any nuclei present at the end control and complete preservation of scaffold architecture.
Static seeding of hMSC has shown lower presence of living cells with their shape rounded as a result of limited adhesion to scaffold, perfusion seeding has shown improved results with more living cells, alligned with the direction of the flow, with star-shape conformation resulting from adhesion to the scaffold.In experimental reconstruction of porcine common bile duct all pigs survived the observation period with one pig developing a bile duct stenosis, similar to the control group. Histological evaluation has shown partial reepithelialization and biointegration into recipients tissues at 4 weeks.
Conclusion: Decellularization and recellularization of porcine eBD with hMSC has been succesfull,
demonstrating crutial role of bioreactors and mechanical stimulation in recellularization process. Theese results showed promissing outcomes as a step towards future in vivo testing of eBD recellularized with hMSC.
Funding: The study was supported by the Charles University, project Grant Agency of Charles
University No. 434522 and Cooperation project “Surgical disciplines” (COOPERATIO-207043,
Charles University).
Study program: Doctoral study –Surgery | Year of study: 2
ID: 1132