Summary: This project explores mechanisms underlying development of seizures in the immediate aftermath of traumatic brain injury (TBI). Early onset seizures are among the most serious morbidities with traumatic TBI. Yet our understanding of the mechanisms that precipitate early seizures is quite incomplete, in part, because most studies report changes in neuronal function when biochemical and molecular studies indicate that significant changes in gene regulation and protein expression have already occurred. To address this gap in our understanding, we modified an in vitro TBI stretch injury model using networks of cultured cortical neurons in which injury is confined to a localized area, but neuron electrical activity can be measured almost immediately. Our novel finding is that hyperexcitability, i.e. dramatically increased spontaneous action potential and bursting activity, is observed within minutes after stretch injury, but only in ?non-injured? neurons located away from the injury site. This hyperexcitability in the non-injured neurons is analogous to activity patterns in in vivo models of TBI where hyperexcitability is thought to precipitate seizure-like discharges. Because hyperexcitability is observed in non- injured neurons only, we hypothesize that reduced inhibitory neurotransmission from injured neurons disinhibits electrical activity in surrounding non-injured neurons. To test this hypothesis, (1) we will determine whether acute hyperexcitability is due to changes in excitatory or inhibitory neurotransmission from injured neurons or due to intrinsic changes in non-injured neurons using electrophysiologic and histologic approaches. (2) We will determine how acute hyperexcitability in non-injured neurons arises from altered dynamics of adjoining injured neurons using genetically-encoded membrane potential sensors to map spatiotemporal changes in electrical activity in physically and functionally defined neurons. These data will be analyzed to determine how stretch injury affects signal propagation and therefore information dynamics in the neural network. Altogether, proposed experiments will allow us to establish a new in vitro model for TBI-induced seizures and, using state-of-the-art molecular techniques, gain an unprecedented understanding of how acute alterations in network function produce hyperexcitability and post-traumatic seizures. In doing so, this project will highlight potential mechanisms to explore in future experiments using in vitro and in vivo TBI models as well as potential approaches to minimize early-onset seizures after TBI. Summary: