Role of P2Y2 in the Regulation of Vascular Tone and Control of Breathing

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PROJECT SUMMARY/ABSTRACT Chemoreception is the mechanism by which the brain regulates breathing in response to changes in tissue CO2/H+. This process is critical for maintaining brain CO2/H+ levels within the narrow range that is conducive for normal neural function. Chemosensitive neurons in the Retrotrapezoid Nucleus (RTN) are directly responsible for increasing respiratory activity and send excitatory projections to respiratory rhythm generating centers in response to CO2/H+. Chemosensitive astrocytes in the RTN release ATP in response to CO2/H+ that directly gains up neuronal activation to modulate respiratory activity and indirectly constrict RTN arterioles by activation of P2Y2/4 receptors. Arteriole vasoconstriction prevents stimulus washout which gains up neuronal and astrocytic responses to CO2/H+. An unexplored area of brainstem chemoreceptor research is the role heterogenetic vasculature in supporting regional functionality. The long-term goal for this fellowship will be to molecularly characterize heterogenetic arteriole cell types in brainstem chemosensing regions and non-chemosensitive areas to provide potential targets for therapeutic action in breathing related disorders. This will be achieved by using cell isolation methods in transgenic fluorescent mouse models for isolation of both endothelial cells and vascular smooth muscle cells. Subsequent FACS and purinergic receptor expression profiling using qPCR will demonstrate the unique properties of arterioles in different brain regions, as previously seen in vitro. By using transgenic knock out mouse models, specific purinergic receptors on arteriole cell types can profiled for functionality and importance for a normal breathing phenotype. This will be accomplished with selective purinergic receptor isoform knock out mouse models using both in vivo and in vitro studies. In vitro arteriole slice recordings compare and contrast arteriole behavior. Whole animal plethysmography and viral knockdown models confirm in vitro arteriole recordings as well as provide evidence on the impact of vasculature dysfunction in whole animal breathing physiology. These results can be utilized further by other researchers in the field for more in-depth investigation of heterogenetic vasculature in other areas of the brain and body as well as provide molecular targets to potentially remediate breathing phenotypes in disease states.