Cocaine-mediated induction of MCP-1 expression involves NF-κB activation. (A) Exposure of BV-2 cells to cocaine resulted in time-dependent increase in phosphorylation of the p65 subunit of NF-κB in the nuclear fraction, with a concomitant decrease in the cytosolic fraction. Reciprocally, cocaine exposure resulted in increased phosphorylation of IκBα in the cytosolic fraction of BV-2 cells. (B) Pretreatment with the IκBα inhibitor SC514 resulted in inhibition of cocaine-mediated induction of MCP-1. (C) Overexpression of the mutant but not the full-length p65/RelA NF-κB construct resulted in abrogation of cocaine-mediated induction of MCP-1. BV-2 cells exposed to cocaine in the presence or absence of σ-1R antagonist BD1047 (D), σ-1R siRNA (E), Src inhibitor PP2 (F), or NADPH inhibitor apocynin (G) were examined for cocaine-mediated translocation of NF-κB. Representative immunoblots and the densitometric analysis of p-P65 NF-κB/histone from 4 separate experiments are presented. All the data are means ± SD of 4 individual experiments. *P < .05; **P < .01 versus control group; #P < .05; ##P < .01 versus cocaine-treated group. (H) Schematic illustration of NF-κB binding consensus sequence on the MCP-1 promoter region. (I) ChIP assay demonstrating cocaine-mediated binding of p65NF-κB to the MCP-1 promoter. The image is representative of 3 independent experiments.