GABAergic INterneurons signaling ImbalancE; A promising target underlying PFC-dependent cognitive flexibility defects.

Project Information

GinieEffect

Grant agreement ID: 101067655

DOI

10.3030/101067655

Project closed

EC signature date 20 June 2022

Start date 1 July 2022

End date 30 June 2024

Funded under

• Marie Skłodowska-Curie Actions (MSCA)

Total cost

No data

EU contribution

€ 153 486,72

Coordinated by

IDRYMA TECHNOLOGIAS KAI EREVNAS
Greece

Periodic Reporting for period 1 - GinieEffect (GABAergic INterneurons signaling ImbalancE; A promising target underlying PFC-dependent cognitive flexibility defects.)

Reporting period: 2022-07-01 to 2024-06-30

Summary of the context and overall objectives of the project

The Covid-19 outbreak has highlighted the significance of adapting our decision-making and behavior in response to unforeseen changes in our surroundings. In vertebrates, prefrontal cortex (PFC) has the cognitive control to modify the animal behavior when the rules change, but the exact mechanisms underlying PFC function at the neuronal level are still largely unknown. Inhibitory GABAergic interneurons (INs) regulate the overall excitability of the prefrontal cortex by reducing the firing rate of the major excitatory pyramidal neurons and thereby fine-tuning the balance between excitation and inhibition. GenieEffect aimed to combine behavioural studies in freely moving mice with neuronal circuit modeling to long-term dysregulate the INs function and investigate their role on the PFC circuit function, with respect to flexible behaviour. A deeper understanding of the role of INs at the neuronal level is important in order to better understand complex behaviors such as decision making. In addition, it can help us better understand traumatic brain injuries, diseases and disorders that involve the PFC region and impact decision-making such as: frontal lobe disorders, executive function disorders, substance use disorders and neurodegenerative diseases. Enriching our basic scientific knowledge on the PFC can help healthcare providers in the future to better understand, diagnose and treat the PFC-dependent psychiatric and neurological disorders.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

GenieEffect’s first two work packages (WP1 and WP2) reflect the two main objectives of the research project. WP1 was dedicated to experimental work and WP2 was dedicated to the modeling of the PFC circuit.
In W1, the objective was to manipulate inhibitory signaling in the mouse prefrontal cortex (PFC) by silencing parvalbumin-positive (PV) inhibitory neurons using genetic techniques. The impact of this manipulation was assessed in a two-choice assay testing flexible behavior. However, the extensive training required for the complex techniques and the need to standardize multiple experimental steps limited the sample size of the mice used, resulting in inconclusive results. Nonetheless, the thorough testing and standardization of the surgical procedures and the behavioral paradigm conducted during this study will significantly benefit future research endeavors.

At the same time, I had the opportunity to standardize an alternative to the typical water deprivation during the days when no behavioral experiments were taking place. On these days water deprivation was replaced with giving citric acid water (first introduced for mice experiments by Urai et al, 2021) to the animal ad libitum. This alteration, while not impending the behavioral training process, ameliorated the animal welfare by helping it maintain a healthier weight, and reduced the labor intensity required from the experimenter on a daily basis. The standardization of this method according to our specific experimental requirements primarily benefited the host lab, while also serving as a valuable example for other research facilities looking to adopt this approach.

During my secondment at Prof. Pitsouli's lab at the University of Cyprus, I received comprehensive training in staining procedures, advanced microscopy techniques, and utilization of relevant software tools. This included proficiency in immunohistochemical staining of tissues with mitosis, nuclear, and tracheal markers, operating the Leica SP8 confocal microscope, and image processing using FIJI (ImageJ) software. Upon my return, I successfully transferred this knowledge to my host lab.

Regarding the WP2, I utilized Dendrify 2.1.4 an open-source Python package developed by a colleague within our lab, to develop a neocortex pyramidal neuron model, that features seven dendritic compartments encompassing dendritic properties. This model can be used to develop an artificial network of this region in order to simulate physiological processes and predict behavioral outcomes.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

Understanding the physiological basis of traumatic brain injuries, diseases and disorders that involve the PFC region and impact decision-making is very crucial in order to develop therapeutic strategies in the future. GenieEffect provides some insights into how to further investigate the role of inhibitory interneurons in the prefrontal cortex region in flexible beaviour using mice.

All the experimental data of GenieEffect are shared with my colleagues in the host lab as they provide valuable insights in refining experimental design practices by offering critical details and enhancements that enrich the existing know-how for the existing surgical and behavioural protocols. The advanced imaging techniques training I received at the secondment lab, along with proficiency in relevant software usage and data processing, has been successfully integrated into the host lab. These acquired skills are now being implemented as new techniques within the host lab's research framework. The computational model I have developed for the primary excitatory pyramidal neuron of the prefrontal cortex holds significant potential for future computational studies. This model has the potential to serve as a foundational element in constructing a prefrontal cortex circuit. The overarching objective would be to use this network to simulate physiological functions within this brain region and manipulate distinct neuronal populations to predict behavioral responses. At a later stage, these predicted behaviors can be validated through in vivo experiments involving mice.

Upon completion of the data analysis, my intention is to disseminate the findings on the Zenodo repository (doi: 10.5281/zenodo.13371667) making them accessible to the wider research community. These data serve as a foundation for future experiments, offering knowledge and valuable insights that can propel further studies. Such studies will advance our understanding of the physiology and pathophysiology of the PFC region and its impact on both normal flexible behavior and related disorders.