Methamphetamine treatment increased the expression of IL-6 and IL-8 in the astrocytes. SVGA astrocytes were grown on coverslips and were treated with 500 μM of methamphetamine (MA) for 24 hours followed by methanol-fixation. Golgi-Stop™ was added 6 hours prior to fixing in order to block the release of the cytokines from the cells into the supernatants. The coverslips were blocked, and then incubated with primary antibody for IL-6/IL-8 and Glial Fibrillary Acidic Protein (GFAP) and secondary antibodies conjugated with Alexafluor488 to detect IL-6 and IL-8 and an antibody conjugated with Alexafluor555 to detect GFAP. The coverslips were then mounted using medium containing DAPI and were observed under a confocal microscope to determine the localization and expression of IL-6 (A) and IL-8 (B). Untreated cells were stained individually for IL-6, IL-8 and GFAP and did not show any non-specific fluorescence (data not shown).