Dynamic temporal regulation of astrocyte coupling to shape neuronal activity during acquired epilepsy development

项目总结

Project Summary Astrocytes couple into networks of hundreds of cells. Impaired astrocyte coupling is associated with epilepsy, but there is no consensus on whether reduced coupling promotes or counteracts abnormal neuronal activity, which precedes seizures. A lack of astrocyte coupling can promote seizures;1 yet, the opposite has also been shown: abnormal neuronal activity and seizures were reduced after coupling was inhibited1-3. An integrated view that accounts for both findings is needed to reveal how astrocyte coupling modulates epilepsy. In acquired epilepsy, which is initiated by a neurological insult such as traumatic brain injury (TBI), many studies have demonstrated dysfunction of gap junctions (GJs) and dysregulation of Connexin43 (Cx43). Cx43 forms GJ responsible for astrocyte coupling leading to the conclusion that reduced coupling may contribute to seizures. Studies in Cx43 knockout mice suggest that the timing and duration of reduced astrocyte coupling may determine if abnormal neuronal activity is promoted or counteracted. This has not been tested due to a lack of tools that dynamically modulate coupling. To reduce or restore astrocyte coupling dynamically, we generated viral constructs that can be induced to express functional or mutated Cx43 for variable durations at different stages of acquired epilepsy. These Cx43 mutants inhibited astrocyte coupling and induced neuronal hyperexcitability in vivo. To assess the relationship between astrocyte coupling and neuronal activity, we will use a model of acquired epilepsy that progresses to spontaneous seizures after mild TBI in the absence of many confounding factors5. This model recapitulates three key aspects of Cx43 pathology: astrocyte coupling is reduced, Cx43 protein is increased and increased phosphorylation at Cx43 serine 368. This post-translational modification alters GJ conductivity, and is associated with internalization of GJ. Yet, the critical upstream signaling causing reduced astrocyte coupling and its effects on neuronal activity during different stages of acquired epilepsy must be revealed as a foundation for future therapeutic targeting. This proposal will generate an integrated model of astrocyte coupling modulation in acquired epilepsy that aims to unify previous findings. Cx43 function will be dynamically manipulated to 1) determine if the timing of reduced astrocyte coupling modulates abnormal neuronal activity 2) identify the signaling cascade controlling reduced astrocyte coupling and 3) determine if and when restoring astrocyte coupling prevents acquired epilepsy. Acquired epilepsy affects 65 million people worldwide and is notoriously difficult to treat. Even after decades of research and the development of new anti-epileptic drugs targeting neurons, one third of patients still suffer from drug-resistant epilepsy. Targeting astrocytic Cx43 might be an option, but the first step towards therapy is determining when reduced coupling is adaptive and when it is maladaptive.