Engagement of σ-1R is critical for cocaine-induced MCP-1 expression in microglia. (A) RNA isolated from cocaine-treated BV-2 cells was subject to RT-PCR analysis using σ-1R primers. (B) Pretreatment of BV-2 cells with σ-1R antagonist BD1047 abolished cocaine-mediated induction of MCP-1 expression in BV-2 cells. (C) Whole-cell lysates from BV-2 cells transfected with either σ-1R or nonsense (Non) siRNAs were subject to Western blot analysis using antibodies specific for σ-1R. (D) σ-1R siRNA, but not Non siRNA, inhibited cocaine-mediated induction of MCP-1 expression. (E) BV-2 cells were treated with cocaine and double-stained using antibodies specific for ganglioside GM1-lipid raft marker (red TRITC fluorescence) or σ-1R (green FITC fluorescence). Overlay images are shown in the right panel. Data are representative from 3 typical experiments. Scale bars all indicate 20 μm. (F) Role of lipid rafts play in cocaine-mediated induction of MCP-1 in BV-2 cells. Cells were pretreated with MβCD (1mM) followed by treatment with cocaine. Supernatant fluids were harvested at 12 hours after cocaine treatment, followed by assessment of MCP-1 expression by ELISA. All the data are presented as means ± SD of 4 individual experiments. *P < .05; **P < .01 versus control group; #P < .05; ##P < .01 versus cocaine-treated group.