Cortical astrocytes are genetically diverse; however, the physiological relevance of this is far from understood because contributions of functionally discrete astrocyte subtypes to whole animal behavior is largely unknown. The most compelling evidence supporting the possibility that astrocytes may have specialized physiological roles is at the level of the brainstem where subsets of astrocytes contribute to the CO2/H+-dependent drive to breathe (respiratory chemoreception) and inspiratory rhythm generation. Therefore, we will use the respiratory circuit as a model system to identify unique genetic profiles of functionally defined subsets of astrocytes. We will use RNA-seq to perform an unbiased evaluation of transcript expression in all cell types isolated from chemosensitive regions, the inspiratory rhythm generating region and non-respiratory brainstem and cortical regions. This analysis will focus on cell populations identified as astrocytes based on expression of cell type specific markers. To correlate gene expression with function, we will use electrophysiological or Ca2+ imaging techniques to functionally identify individual astrocytes in slices from each region for subsequent harvest and targeted qPCR for genes expressed by each astrocyte subpopulation. The proposed work will enable development of tools to study contributions of discrete subpopulations of astrocyte to respiratory behavior. Also, since disruption of astrocyte CO2/H+ chemoreception contributes to disordered breathing in certain diseases (e.g. Rett syndrome), the ability to selectively target CO2/H+-sensitive astrocytes would be an attractive strategy for treating disordered breathing associated with this condition.
Recent evidence suggests that subsets of astrocytes in key brainstem respiratory control centers contribute to respiratory behavior. However, nothing is known regarding the molecular identity of functionally discrete subsets of astrocytes. The proposed work will use a novel combination of electrophysiology and gene expression analysis to characterize the gene expression profile of functionally defined subsets of astrocytes across multiple levels of the respiratory circuit.
