Nuclear factor kappa B in the dorsal raphe of macaques: An anatomical link for steroids, cytokines and serotonin

Objective: Nuclear factor kappa B (NFκB) is a transcription factor that activates gene expression in response to proinflammatory cytokines, and elevated cytokines are associated with depression, which has a serotonergic component. We questioned (1) whether serotonin neurons contain NFκB, (2) whether NFκB detection with immunocytochemistry is changed in the dorsal raphe nucleus (DRN) by ovarian hormone treatment and (3) whether ovarian hormones regulate midbrain NκB gene or protein expression. Methods: Monkeys were spayed and treated with placebo, estrogen (E), progesterone (P) or E+P for 1 month (n = 4 animals/treatment group), and the midbrain was harvested for immunocytochemistry and stereology. An antibody that detects nuclear location-specific (NLS)-NFκB p65 was applied, and the numbers of NLS-NFκB-immunopositive cells were counted in 9 sections of the DRN. Additional monkeys were used for Western blot analysis and quantitative reverse transcription-polymerase chain reaction (RT-PCR) for NFκB p65. Results: In placebo-treated macaques, neurons were double-immunostained for serotonin and nuclear NFκB p65 throughout the DRN. The mean total number of NFκB-positive cells equalled 2178 (and standard error of the mean [SEM] 129) in the placebo group, 1631 (SEM 221) in the E-treated group, 2314 (SEM 186) in the P-treated group and 1162 (SEM 100) in the E+P-treated group (analysis of variance p = 0.003). The E-treated and E+P-treated groups had a significantly lower density of cells stained positive for NFκB than the placebo or P-treated groups (post hoc). Unmasking of NLS-NFκB immunostaining in the DRN revealed dense immunostaining in the cytoplasm of large dorsal raphe neurons. There was no difference between treatment groups in the amount of NFκB p65 detected by Western blot or in the relative expression of NFκB p65 mRNA with quantitative RT-PCR. Conclusions: These observations are consistent with the notion that gene and protein expression of NFκB are constitutive but that ovarian hormones can decrease the nuclear location of NFκB in dorsal raphe neurons and, thereby, decrease the ability of NFκB to drive gene expression in response to cytokines.