Figure 5.

Protective effect of NT on glutamate excitotoxicity-induced death of PC-12 cells. (a) Dose-dependent cell viability of PC-12 cell following incubation with varying concentrations of glutamate (0, 1, 5, 10, 15, and 20 mM) for 24 hr. ***p < .001 versus cells untreated with glutamate. (b) Dose-dependent cell viability of PC-12 cell following incubation with varying concentrations of NT (0, 2.5, 5, 50, and 100 µM) for 24 hr. ***p < .001 versus cells untreated with NT. As revealed in (a) and (b), it was determined that the median lethal dose (LD₅₀) of glutamate and maximum allowable concentration of NT are 15 mM and 10 μM, respectively; thus, those concentrations were selected as the optimal doses for evoking glutamate excitotoxicity and applied for all the subsequent in vitro studies. In this regard, each PC-12 cell group treated with vehicle, 15 mM glutamate, or 15 mM glutamate plus 10 μM NT will be referred to as control, GLU, or GLU+NT groups, respectively. (c) Cell viability of control, GLU, and GLU+NT groups tested at 24 hr after the different treatments. All bar graphs represent mean ± standard deviation in at least three independent experiments (n = 8 wells per group). ***p < .001 versus control; ^###p < .001 versus GLU group. (d) Representative cell images captured using an inverted microscope for the different groups. Membrane blebbings are depicted with a white arrowhead. Scale bar = 25 μm. GLU = glutamate; NT = nortriptyline.