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Show Visualization of ATP in End Foot Processes of Sustentacular Cells in Mouse Olfactory Epithelium Sara Hogan, Coilccn Hegg, and Associate Professor Mary Lucero Department of Physiology Kodcnts can be used as a M for Human Olfaction Olfactory Epithelium was studied by making coronal 1 sections through the mouse nose Purinergic Receptors are Present in the Olfactory Epithelium In Vivo hxpcriments Visualization of ATP Stores with Quinacrine In Vivo Experiments High Magnification confirms ATP stores in ORNs and basal cells In Vitro Experiments Quinacrine Labeling of ATP in Olfactory Epithelium Role of ATP during damage to Olfactory Epithelium *. 4, In the mammalian olfactory epithelium, sustentacular support cells extend from the apical surface to the basal lamina where their end foot processes come in close contact with olfactory stem cells. Our hypothesis is that sustentacular cells play a critical role in injury-evoked olfactory neuron regeneration. We postulate that when ol-factotoxicants damage cells of the olfactory epithelium, ATP is released and activates P2Y purinergic receptors on sustentacular cells, thereby eliciting increases in intracellular calcium and resulting in release of neurotrophic fac-tors and ATP from vesicles in sustentacular cells. As a first step in testing this hypothesis, we determined that ATP is present in end foot vesicles by using the fluorescent drug quinacrine to detect the presence of vesicular ATP in mouse olfactory epithelium. Postnatal day 3 mice were injected intraperitoneally with 0 or 50 u.g/g quinacrine. One hour later they were anesthetized, decapitated, flash frozen, embedded in freezing media and cryosec-tioned. Tissue sections (12 u.m) were immediately examined for quinacrine fluorescence. Punctate staining was predominantly observed in the basal stem cell layer of the 50 u.g/g quinacrine group, although puncta also appeared in the middle zone of the olfactory epithelium. In contrast, very little fluorescence was observed in the sham control. This data suggests that ATP is located in vesicles predominantly located in the basal cell layer. In addition, we incubated live olfactory epithelium slices (300 u.m) in 5 uM quinacrine for 30 minutes and observed punctate fluorescence in the basal zone of the olfactory epithelium. Our goal is to visualize the P2Y-mediated decrease in quinacrine fluorescence as an assay for determining the pathways involved in signaling damage and for initiating regeneration in the olfactory epithelium. |