Figure 3.

(A) Time dependent effect of Cya-EC on ³⁵S-cystine uptake in HT22 cells. Cells were treated with 25 μM ³⁵S-cystine alone or in the presence of 10 μM Cya-EC. After solubilization of the cells in 0.2 M NaOH, aliquots were taken for scintillation counting and protein determination. Results are presented as cpm taken up per μg cellular protein. Similar results were obtained in three independent experiments. (B) Dose dependent effect of Cya-EC on ³⁵S-cystine uptake in HT22 cells. Cells were treated for 50 min with 25 μM ³⁵S-cystine alone or in the presence of increasing concentrations of Cya-EC. After solubilization of the cells in 0.2 M NaOH, aliquots were taken for scintillation counting and protein determination. Results are presented as cpm taken up per μg cellular protein. Similar results were obtained in three independent experiments. (C) Correlation between the dose dependent effects of Cya-EC on ³⁵S-cystine uptake and GSH maintenance. (D) Effect of pretreatment time on the Cya-EC-mediated increase in ³⁵S-cystine uptake. Cya-EC was added to ³⁵S-cystine immediately before addition to the cells or incubated with ³⁵S-cystine at 37^oC for 10-50 min before addition to the cells for 10 min. After solubilization of the cells in 0.2 M NaOH, aliquots were taken for scintillation counting and protein determination. Results are the average ± SEM of three independent experiments. (E) Cya-EC does not protect from cystine deprivation. HT22 cells were treated with 5 mM glutamate or cystine-free DME alone or in the presence of 10 μM Cya-EC. 24 hr later cell survival was measured by the MTT assay. The results are the average ± SEM of three independent experiments.