OSMOSENSITIVE RELEASE OF TAURINE AND GLUTAMATE PARTICIPATE IN CELL VOLUME REGULATION IN RETINAL MÜLLER CELLS

VANINA NETTI; MARTHA PEREZ DOMINGUEZ; HERMINIA PASANTES MORALES; GERARDO RAMOS MANDUJANO; ALEJANDRO PIZZONI; CLAUDIA CAPURRO

Rio de Janeiro

Congreso; International Union of Pysiological Sciences (IUPS) 38th World Congress; 2017

International Union of Pysiological Sciences

Müller cells are involved in controlling extracellular homeostasis in the retina, where neural activity alters osmotic gradients, favoring cell swelling. This swelling is followed by a regulatory volume decrease (RVD) mediated by an efflux of osmolytes and water transport by Aquaporin-4 (AQP4). We previouslydemonstrated in a human Müller cells (MIO-M1) that RVD partially depends on an efflux of KCl, but an important fraction of this response may depend on the release of other solutes, such as Taurine (Tau) and Glutamate (Glu), poorly studied in Müller cells. Thus, the aim of this study was to evaluate the contribution of Tau and Glu efflux to RVD and to identify the involved signaling pathways in MIO-M1 cells. Cell volume was evaluated by FURA-2 AM fluorescence and [3H]-Tau/[3H]-Glu release was assessed by measuring radioactivity in culture supernatant after the hypotonic shock (ΔOsm:100 mOsm). Results showed that MIOM1 cells exposed to hypotonicity exhibited an efflux of [3H]-Tau and [3H]-Glu which was prevented by the volume-regulated anion channel (VRAC) blocker DCPIB, reducing RVD by 60%. During hypotonicity,[3H]-Tau efflux was decreased in Ca2+-free medium and even further reduced by the intracellular Ca2+ chelator BAPTA-AM, whereas [3H]-Glu release was insensitive to both conditions. RVD was unaffected in Ca2+-free medium but it was lowered by 50% in the presence of BAPTA-AM. PI3K inhibition with Wortmannin reduced both [3H]-Tau and [3H]-Glu efflux by 47 and 75%, respectively, decreasing RVD. On the other hand, PKC inhibition with Gö6976 did not affect [3H]-Tau and [3H]-Glu efflux, but RVD andosmotic water permeability were increased by 75% and 25%, respectively. These results suggest that Tau and Glu released by VRAC have a relevant role in RVD in MIO-M1 cells. PI3K activation represents a major regulatory mechanism for both osmolytes efflux, but only Tau release was influenced by Ca2+. Ca2+-independent Glu efflux may support RVD in Ca2+-free medium, but it may not be sufficient when Ca2+ release from intracellular stores is impaired. PKC is not directly involved in modulating osmolytes efflux, but it may affect water permeability mediated by AQP4, decreasing the RVD response. These findings support a role of Tau and Glu as osmolytes for RVD in Müller cells, key regulators of the retinalmicroenvironment.