Stimulation of EAAC1 in C6 glioma cells by store-operated calcium influx

Abstract

This study investigated how modulation of intracellular calcium alters the functional activity of the EAAC1 glutamate transporter in C6 glioma cells. Pre-incubation of C6 glioma cells with the endoplasmic reticulum Ca2+ ATP pump inhibitor, thapsigargin (10 microM) produced a time-dependent increase in the Vmax for D-[3H]aspartate transport that reached a maximum at 15 min (143% of control; P<0.001) that was accompanied by increased plasma membrane expression of EAAC1 and was blocked by inhibition of protein kinase C. Pre-incubation of C6 glioma cells with phorbol myristate-3-acetate (100 nM for 20 min) also caused a significant increase in the Vmax of sodium-dependent D-[3H]aspartate transport (190% of control; P<0.01). In contrast, in the absence of extracellular calcium, thapsigargin caused a significant inhibition in D-[3H]aspartate transport that was not mediated by protein kinase C. Blockade of store-operated calcium channels with 2-aminoethoxydiphenyl borate (50 microM) or SKF 96365 (10 microM) caused a net inhibition of D-[3H]aspartate uptake. Co-incubation of C6 glioma cells with both thapsigargin and 2-aminoethoxydiphenyl borate (but not SKF 96365) prevented the increase in D-[3H]aspartate transport that was observed in the presence of thapsigargin alone. Furthermore, 2-aminoethoxydiphenyl borate, but not SKF 96365, reduced the increase in intracellular calcium that occurred following pre-incubation of the cells with thapsigargin. It is concluded that, in C6 glioma cells, stimulation of EAAC1-mediated glutamate transport by thapsigargin is dependent on entry of calcium via the NSCC-1 subtype of store operated calcium channel and is mediated by protein kinase C. In contrast, in the absence of store operated calcium entry, thapsigargin inhibits transport.

Two new coordination polymers containing dicyanidoargentate(I) and dicyanidoaurate(I): synthesis and characterization, and a detailed in vitro investigation of their anticancer activities on some cancer cell lines

“…Moreover, all cell lines treated with C1 and C2 showed apoptotic indicators, which was confirmed by May-Grunwald-Giemsa stain (Figure 8). These results were somewhat similar to those of previous studies by Hall et al, 45 Rackham et al, 46 Thati et al, 47 Wang et al, 48 Wang et al, 49 and Zhu et al 50 Thus, based on observations in the literature, the appearance of cell lines treated with C1 and C2 clearly indicated the antiproliferative and low-cytotoxic effect of these complexes. Moreover, the results of DNA fragmentation assays and TUNEL assay indicate that these novel complexes may inhibit cell proliferation through the induction of apoptosis.…”

Muscarinic Receptor Stimulation of d-Aspartate Uptake into Human SH-SY5Y Neuroblastoma Cells Is Attenuated by Hypoosmolarity

“…9). These findings are consistent with results recently obtained for C 6 glioma cells in which a sustained entry of Ca 2ϩ , elicited independently of receptor activation, resulted in a PKC-dependent increase in both the cell-surface expression and activity of EAAT3 (Murphy et al, 2009). Optimal Daspartate transport in SH-SY5Y cells under both basal-and Oxo-M-stimulated conditions also requires the maintenance of cytoskeletal integrity (Fig.…”

Bone cell elasticity and morphology changes during the cell cycle

“…Prior to experiments, culture medium was removed and cells were washed in Krebs bicarbonate buffer (KBB) (Murphy et al, 2009). The slide to which the cells were adhered was positioned on the AFM stage and cells remained immersed in KBB for the duration of experiments.…”