Development of a novel NPY-sensitive microelectrode to detect endogenous NPY release - the main objective of this proposal is to develop an NPY-sensitive microelectrode for continuous measurement of…

Development of a novel NPY-sensitive microelectrode to detect endogenous NPY release - the main objective of this proposal is to develop an NPY-sensitive microelectrode for continuous measurement of NPY using an aptamer-modified platinum microelectrode with a total size of less than 25 ΜM. This will allow the research community to monitor endogenous NPY release in the intricate microenvironment of brain slices and single cells. In this way, we can establish connections between NPY levels and neurological disorders. To accomplish this, we propose a strategy for enhancing the detection of NPY by our previously developed NPY biosensor. Our initial findings indicate that our NPY-sensitive microelectrode can measure NPY with a sub-second time resolution, collecting 500 data points using intermittent pulse amperometry (IPA). By utilizing the temporal precision offered by IPA and distinguishing non-faradaic currents and the decay of faradaic electrical currents, our objective is to optimize the analytical signal of NPY measurements to improve sensitivity and selectivity at the same temporal resolution of 500 data points per second. As part of our efforts to understand the biological role of NPY, single-cell measurements will be performed to detect the endogenous release of NPY and comprehend its effects at a cellular level. Until now, measurements have not been possible, and given the significant importance of NPY release in neuronal functions, we present our NPY-sensitive microelectrodes as a tool to improve our understanding of the NPY effects in biological systems. Based on our preliminary work, our NPY biosensor has the ability to measure dynamic NPY concentrations as low as 2 PG/ML. To achieve better biosensor performance and understand the parameters that affect IT, IT is crucial to conduct thorough testing on several NPY-releasing cell lines that provide fast NPY release. This will be accompanied by iterative optimization of procedures and data processing. In addition, we intend to perform a comparative analysis of the optimized NPY biosensor compared to our existing NPY biosensor. At the same time, we will continue improving on our endogenous NPY measurements to detect NPY released endogenously in acute hippocampal slices using the proposed electrochemical approach, which has not been done before. We will compare the release of endogenous NPY by activating NPY cells via optogenetics or electrical stimulation with a physiological activity pattern. This comparison will be done in slices from mice with different levels of NPY expression and across several hippocampus pathways. The NPY biosensor microelectrodes created in this research are seen as an essential instrument that connects the NPY release, circuit function, and behavior. These techniques can be applied to different neuropeptides, offering a promising opportunity for a new age in measuring the dynamics of neuropeptide brain chemistry.