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
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.
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.