Fig. 2

HIV-tat increased mRNA and Cx43 protein expression as well as gap junctional communication in human primary astrocytes. Human primary astrocytes were treated with recombinant HIV-tat protein (100 ng/ml), and expression and function of Cx43-containing channels were analyzed. a Staining for the nucleus (DAPI, blue staining, in the insets in the left column), Cx43 (Alexa 488, green staining), and GFAP (an astrocyte marker, Cy3, red staining) in untreated (control) and HIV-tat-treated conditions (HIV-tat) after 24 h of treatment. The last panel represents the merge of all colors. Bar 75 μm. Arrows denote gap junction plaques. b qRT-PCR for Cx43 and GAPDH mRNA using untreated (control) and HIV-tat-treated cultures of astrocytes at different time points (0, 6, 12, 24, and 48 h). No significant differences in mRNA Cx43 expression were detected in control cells (white bars). HIV-tat treatment increased Cx43 mRNA expression in a time-dependent manner (*p ≤ 0.001, n = 4, black bars). c Western blot analysis of Cx43 protein expression in control and HIV-tat-treated human astrocytes for 6, 12, 24, and 48 h. As a loading control, tubulin was used (tub). As a positive control for Cx43 and its phosphorylation, mouse astrocytes were used (+). d Dye coupling experiments using Lucifer Yellow (LY) showed that in both control and HIV-tat-treated cultures, dye spread into neighboring cells was 100 % (images not shown). However, HIV-tat increased the numbers of coupled astrocytes for each microinjection (*p ≤ 0.003, n = 4), indicating increased gap junctional communication