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Show RESEARCH POSTERS ON THE HILL SPRING 2013 CHARACTERIZING THE PROPERTIES OF CELL VOLUME REGULATION IN RETINAL NEURONS AND GLIA A ROLE FOR THE MECHANOSENSITIVE CATION CHANNEL TRPV4 Andrew Jo1, Daniel A. Ryskamp' ', David Knzaj' • 'Department of Ophthalmology and Visual Sciences. John A. Moran Eye Center, 'Department of Physiology, and 'Interdepartmental Program in Neurosaence University of Utah School of Medicine, Salt Lake City, UT 84132. u THE UNIVERSITY OF UTAH TRPV4 is expressed in R G Cs l RGC and M C response to osmotic pressure , =^M.. tore. (B) g " MOIkr glia (A) and R G Cs (B) thai were inhibited by HC.a rRPV4aj ia Stretch-induced TRPV4-mediated calcium influx exacerbates the swelling extent in hypotonicity by actin de polymerization CHARACTERIZING THE PROPERTIES OF CELL VOLUME REGULATION RETINAL NEURONS AND GLIA: A ROLE FOR THE MECHANOSENSITIVE CATION CHANNELTRPV4 Andrew Jo (David Krizaj, Daniel Ryskamp) Department of Ophthalmology and Visual Sciences University of Utah All cells, including retinal neurons and glia, must sense and adapt to physical changes in their local environment (e.g. changes in osmotic pressure). Osmotic water flux can cause aberrant cell volume changes, which can contribute to tissue damage, edema, and neuronal hyperexcitability and excitotoxicity. W e hypothesized that force-sensitive proteins enable retinal cells to monitor their physical form (e.g. volume) and help maintain homeostasis by regulating cell volume. To test this, w e first investigated the properties of cell morphology when cells were bathed in solutions with different tonicities. Under these conditions, w e measured changes in cell volume due to osmosis in retinal ganglion cells (RGCs) and Muller glia. W e found that RGCs exhibit relatively less swelling than Muller glia in hypotonic saline. RGCs were unable to actively adjust their volume, whereas Muller glia reduced their swollen volume in the continued presence of hypotonicity. The regulation of cellular volume often involves calcium signals. W e tested whether calcium plays a role in the regulation of retinal cell volume. Free calcium within the cells was sequestered using cytosolic BAPTA, which decreased the extent of hypotonic swelling. This demonstrates that calcium elevations increase the extent of cell swelling. Because cell volume was dependent on calcium, which was elevated by membrane stretch, w e hypothesized that the osmosensitive cation channel TRPV4 would transduce osmotic pressure and contribute to cell volume regulation. In an experiment using a selectiveTRPV4 inhibitor, the extent of hypotonic-induced swelling was reduced. Thus, the opening of TRPV4 leads to a calcium influx that exacerbated cellular swelling. Finally, w e tested the idea that excessive calcium influx (calcium overload) induced by stretch-activated channels such as TRPV4 might compromise cellular viability. W e used TUNEL and Live/Dead assays to observe that indeed, hypotonicity induced RGC apoptosis/death in retinal wholemount preparations.Thus, inhibition of TRPV4 might alleviate the deleterious effects of volume changes in pathological contexts. Our findings therefore have implications for our understanding of retinal mechanotransduction and osmoregulation as well as provide a mechanistic framework for developing new therapeutic strategies aimed at blinding conditions that involve mechanical stress and cellular morphology changes such as ischemia, neuromyelitis optica, macular edema and glaucoma. |
