Figure 3
Met and TQ prevented ethanol destabilization of mitochondrial membrane potential. (A) Flow cytometric analysis of mitochondrial membrane potential (ΔψM) was made with JC-1. Mitochondrial polarization was monitored by flow cytometric analysis of JC-1 stained cells that were cotreated for 12 h with ethanol (EtOH, 100 mM), TQ (25 μM) plus ethanol (TQ+EtOH), Met (10 mM) plus ethanol (Met+EtOH), TQ plus Met plus ethanol (TQ+Met+EtOH) and untreated (Control). A representative collapse in ΔψM is associated with high FL1 fluorescence (green) and low FL2 fluorescence (red). The number in each quadrant indicates cell population in that quadrant as the percentage of total cell population. Loss of ΔψM was associated with an increase in FL1 fluorescence. Quantification of cells with ΔψM (as the percentage of total cell population) induced by ethanol, TQ and Met in different combinations as detected by flow cytometry. Data are the mean ± SEM of three independent experiments (n = 3). The details of procedures are mentioned in materials and methods section. (B) Fluorescence analysis of neurodegeneration in primary cultures of fetal rat brain cortical neurons. Cultures were exposed to growth medium (Control) and with ethanol (EtOH, 100 mM), TQ (25 μM) plus ethanol (TQ+EtOH), Met (10 mM) plus ethanol (Met+EtOH), TQ plus Met plus ethanol (TQ+Met+EtOH) supplements for 12 h before staining with Fluoro-Jade B (FJB; green) and propidium iodide (PI; red). Confocal micrographs of Fluoro-jade-B and PI staining show neurodegeneration in primary cortical neurons. Magnification with 40× objective field, Scale bar = 20 μm.