Long‐term in vivo monitoring of transcription factor (TF) activation in cardiovascular (CV) control regions of mouse brain using bioluminescence imaging (BLI)

Abstract

Many CV diseases involve altered CNS pathways that lead to sustained autonomic dysfunction. The molecular basis of these long-term changes is unknown, but activation of TFs such as nuclear factor kappa-B (NF-κB) and activator protein-1 (AP-1) is likely involved. Conventional assays are not suitable for tracking TF activation longitudinally, especially when tissue is limited. We hypothesized that in vivo BLI would provide a novel means for quantifying patterns of TF activation over time in mouse CV brain sites. C57 mice underwent forebrain CV site gene transfer of adenoviruses (Ad) encoding the firefly luciferase (Luc) gene downstream of response elements for NF-κB (AdNF-κB-Luc, n=7) or AP-1 (AdAP-1-Luc, n=4). Luminescence was quantified (photons/sec) intermittently for 4 wks using Xenogen IVIS 200. Basal TF activity was low but stable in the forebrain for at least 4 wks (AdNF-κB-Luc 10317 ± 1518; AdAP-1-Luc 4178 ± 798 p/s). Systemic challenge at 4 wks with lipopolysaccharide (LPS, 8 μg/g, ip) caused robust activation of both NF-κB (766347 ± 237326, p<0.05) and AP-1 (616510 ± 123474, p<0.05) in this brain region. Co-gene transfer of the suppressor mutant of NF-κB (AdIkB32/36S/A) abolished LPS-induced NF-κB activation (15042 ± 1415, p<0.05), confirming specificity. These data demonstrate the utility and power of BLI for longitudinal quantification of TF activation in CV regulatory centers of the mouse brain in vivo.