SOCE is modulated by AQP2 and TRPV4 affecting both Ca2+ influx and clearance in renal cells.

DI GIUSTO GISELA; PIZZONI ALEJANDRO; FORD PAULA; RIVAROLA VALERIA; LÓPEZ GONZALEZ MACARENA; CAPURRO CLAUDIA

Tucuman

Congreso; . III LAFEBS / IX IberoAmerican Congress of Biophysics / XLV Reunión Anual SAB 2016; 2016

III Latin American Federation of Biophysical Societies (LAFEBS)

Store-operated Ca2+ entry (SOCE) isa homeostatic process regulated by the filling state of the intracellular Ca2+stores.This mechanism is strongly influenced by changes of the driving force for Ca2+influx. In addition, SOCE is modulated through the plasma membrane Ca2+ATPase (PMCA)-mediated cytosolic Ca2+[Ca2+]iclearance with important consequences in downstream signaling pathways1.Ina previous communication we described that AQP2 can modulate SOCE byinteracting with TRPV4, affecting K+ channels and in consequencemembrane potential (Vm). The aim of thepresent work was to further characterize if SOCE modulation by AQP2 depends onchanges of TRPV4 localization, the driving force for Ca2+ entry andCa2+ clearance rates. We used two renal cell lines; one notexpressing AQPs (WT-RCCD1) and another one transfected with AQP2(AQP2-RCCD1). We performed immunofluorescence studies to determineif Ca2+store depletion with thapsigargin (1 μM) can changesubcellular TRPV4 localization. Also, FURA-2 and DIBAC4(3) dyes wereused to monitor [Ca2+]i  and Vm changesrespectively. We found that after Ca2+store depletion, TRPV4 wasenriched only in the membrane of cells expressing AQP2. In addition, SOCEsensitivity to changes in the driving force was evaluated by varyingsequentially either the electrical (high extracellular K+ medium, HK) or thechemical (low extracellular Ca2+ medium, LC) driving force for short periods inthe same cells. WT cells responded strongly reducing [Ca2+]Ibut surprisingly, AQP2 cells showed a minor reduction on [Ca2+]i upondepolarization following introduction of HK medium. Contrarily, both cell linesrespond similarly to LC medium. Experiments designed to evaluate Ca2+clearance revealed that AQP2 expressing cells remove Ca2+ from citosol slowerthan WT cells. These results suggest that AQP2 can modulate SOCE by influencingeither Ca2+ influx and Ca2+ clearance from cytosol.