Objectives: 

Selenium compounds are able to open Ca2+ release channels at the sarcoendoplasmic reticulum membrane, leading to an increase in cytosolic free Ca2+ concentration ([Ca2+]c). Our objective was to investigate the effect of the seleno-organic compound and radical scavenger ebselen on rat hippocampal astrocytes in primary culture.

Materials: 

We have studied cell proliferation and cytotoxicity in the presence of different concentrations of ebselen (1-100 mM), using Alamar-Blue®. We have also employed single cell imaging to monitor changes in [Ca2+]c in fura-2-loaded cells, and confocal laser scanning microscopy to monitor changes in mitochondrial Ca2+ concentration ([Ca2+]m), mitochondrial membrane potential (MMP) and mitochondrial membrane permeability, by loading the cells with rhod-2, TMRM or MitoTracker Green FM, respectively.

Results: 

Our results show that stimulation of astrocytes with ebselen (1-100 mM) dose-dependently reduced cell viability. It also induced a sustained increase in [Ca2+]c, that was higher with increasing concentrations of ebselen. This increase in [Ca2+]c was accompanied by a loss of cell membrane integrity, that was delayed in the absence of extracellular Ca2+ (medium containing 0.5 mM EGTA) and in the presence of the intracellular Ca2+ chelator BAPTA. The antioxidant (100 mM) led to Ca2+ accumulation in mitocondria. Ebselen (100 mM) also induced depolarization of MMP. In addition, in the presence of ebselen (100 mM) we observed a disruption of mitochondrial network, a loss of MitoTracker Green FM content and a diffusion of the dye into the cytosol.

Conclusions: 

Collectively, these data support the likelihood that ebselen, depending on the concentration, reduces viability of rat hippocampal astrocytes via cytosolic Ca2+ accumulation and by its action on mitochondrial activity. Thus, we consider that careful attention should be paid when employing ebselen as a prophylactic agent against brain damage.

Funding sources: Plan Regional Investigación Sanitaria (PRIS10014), Junta de Extremadura-FEDER (GR10010), and PSC was granted a fellowship from Junta de Extremadura (Consejería de Economía, Comercio e Innovación) and European Social Fund.