Candidate-based and Discovery-based Analysis of Astrocytes and Glutamate Neurotransmission

Washington University, Saint Louis MO

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Abstract

? DESCRIPTION (provided by applicant): This proposal explores how neuron-astrocyte interactions facilitate normal neuronal function, and the consequences of removing local astrocytic support while leaving all other conditions unaltered. Astrocytes have proven to be important for promoting normal neuronal function and are also implicated in neuropsychiatric disorders such as mood disorders, schizophrenia, and forms of autism. While we have learned a great deal about this relationship over the past decade, we are confronted with technical issues that impede future progress. The inability to remove astrocytes from a neuronal circuit without changing many additional variables profoundly inhibits our ability to explore this relationship completely. My proposed research overcomes this issue by introducing a novel cell culture system where local astrocyte support is absent on some microislands, while neighboring microcultures contain astrocyte beds. Our preliminary data show that local astrocyte absence results in dramatic, large-scale asynchronous glutamate release but no change in GABA release. Preliminary data also suggests that this is due to axonal, and not somatic or dendritic, deficits in conditions where local astrocyte support is removed. In this proposal I will test the hypothesis that an absence of local astrocytic support modulates action potential propagation in hippocampal pyramidal neurons by altering axon anatomy and/or by influencing axonal potassium channel expression, localization or function. This proposal will test these possibilities with the use of electrophysiology, immunolabeling, and confocal imaging, while more expansively surveying differences between neurons grown with and without astrocytic support through a discovery-based, hypothesis-generating transcriptome analysis, RNA-seq. We expect this approach to uncover networks of transcripts that participate in the observed axonal defects, but we also expect to reveal additional unappreciated roles of astrocytes that remain veiled by traditional candidate- based approaches.