Project Summary The medial entorhinal cortex (mEC) plays a vital role in spatial navigation, learning, and memory. Many neurons in layer II/III of the mEC exhibit spatially tuned firing rates that generate a grid-like (‘grid cells’) pattern when traversing an open field. Grid cell firing rates are modulated by a theta (4-12 Hz) frequency, network-wide oscillation generated via input from the medial septum. Further, higher frequency gamma (40-140 Hz) oscillations are nested within the slower- wave theta oscillation and are believed to help synchronize grid cell spike output. Several studies have demonstrated that grid cells are largely connected through a dense network of fast-spiking interneurons which are critical for the generation of gamma oscillations. However, the functional network connectivity between putative grid cells and fast-spiking interneurons in the mEC, which generate theta-nested oscillations during spatial navigation, are not fully understood. Whole-cell patch clamp recordings remain the standard for measuring intracellular membrane voltage and current, but this technique has relatively low throughput. Recent advances in fluorescent voltage indicators have enabled the imaging of both action potentials and subthreshold activity from tens of neurons during optogenetic stimulation. We propose utilizing the sensitivity of whole-cell voltage clamp recordings to capture network synaptic activity in stellate, pyramidal and fast-spiking interneurons during optogenetic stimulation of different local excitatory and inhibitory cell populations. Following this, we will determine the spike timing of excitatory and inhibitory neurons relative to theta-nested gamma oscillations in the local field potential by imaging intracellular voltage in a densely labeled population during optogenetic stimulation of local excitatory neurons. The combination of these techniques will establish the functional input/output of each cell type necessary for developing and testing potential canonical models of grid cell activity and network synchrony during spatial navigation. The proposed research will be conducted at Boston University in the Rajen Center for Integrated Life Sciences which is home to a multidisciplinary community of neuroscience investigators. This institute combines experts from the Center for Systems Neuroscience and the Neurophotonics Center which fosters a diverse collaborative environment to tackle challenging research projects. Further, the development of my academic training, technical skills, scientific communication, professional skills, and consistent mentorship will ensure the success of this project.