Fig. 2

Massive increase of immune cell trafficking into the prefrontal cortex of the CNS of mice treated with morphine and HIV-1 Tat and systematically infected with S. pneumoniae at the 5th day of post-pellet implantation. a Splenocyte homing was assessed following the adoptive transfer of luciferase-expressing immune cells (1 × 10⁷) into B6CBAF1 recipient mice. The recipient mice were imaged for whole-body bioluminescence at 24 h following the transfer of luciferase-positive immune cells. b The mean bioluminescent signal was measured as total photon emission using Igor image analysis software as explained in Fig. 1b. c Immune cell homing in the perfused brain was assessed and d measured as explained in Fig. 1. e Luciferase activity in the brain lysate was measured (indicative of the number of immune cells) and is presented as histograms of six different animals from each group. Relative luminescence units (RLU) were normalized with the Renilla luciferase values. Error bars indicate the standard error of the mean (SEM) of six different animals from each group (n = 6). Pseudocolor scales are shown to right of the figures. f Confocal microscopy and z-stacked images showing CD45+ immune cells (Alexa Fluor 488-conjugated anti-mouse IgG; green) in the prefrontal cortex brain tissue with S. pneumoniae (Rhodamine Red goat anti-rabbit IgG; red) and counterstained nuclei with DAPI (blue), suggesting cellular mode for bacterial translocation into the CNS of mice treated with morphine and HIV-1 Tat and co-infected with S. pneumoniae. Merged images show co-localization of CD45+ immune cells and S. pneumoniae. Magnification ×600 (upper panel) and ×2400 (lower panel). PP placebo pellet, MP morphine pellet. Scale bar 10 μm. *P < 0.05