SELECTIVE INHIBITION OF THE Na, K–ATPASE ALPHA 2 ISOFORM ALTERS CAPACITATIVE CALCIUM ENTRY IN OPTIC NERVE ASTROCYTES

Abstract

Abstract: : Purpose: The Na,K–ATPase α2 isoform in astrocytes is localized in a pattern that suggests a possible link to endoplasmic reticulum (ER) function. Here we tested whether selective inhibition of Na,K–ATPase α2 alters capacitative calcium entry (CCE), the mechanism by which ER calcium stores refill following emptying. Methods: Rat optic nerve astrocytes were established in primary culture. Na,K–ATPase α2 was selectively inhibited using a low concentration of ouabain. Cytosolic calcium and sodium concentrations were measured using Fura–2 and SBFI respectively. Results: The magnitude of the increase in cytosolic calcium concentration during CCE was almost doubled in astrocytes exposed to 1µM ouabain. The same concentration of ouabain was not sufficient to cause a cell–wide increase of cytosolic sodium concentration, nor to inhibit activity of the rat Na,K–ATPase α1 isoform. To examine ER calcium store size, cells were exposed to the calcium–mobilizing agonist ATP (50µM) in the absence of extracellular calcium. In cells exposed to 1µM ouabain, the calcium response to ATP was increased by ∼80%. Conclusions: The results suggest that selective inhibition of the Na,K–ATPase α2 isoform has the potential to change CCE–mediated calcium entry, alter calcium signaling responses and possibly increase calcium ER store size. The findings may signify interaction and co–localization of store operated calcium channels, the Na–Ca exchanger and Na,K–ATPase α2. .

Keywords: NaK ATPase • calcium • astrocytes: optic nerve head