Responses of perivascular macrophages to circulating lipopolysaccharides in the subfornical organ with special reference to endotoxin tolerance

Circulating endotoxins including lipopolysaccharides (LPS) cause brain responses such as fever and decrease of food and water intake, while pre-injection of endotoxins attenuates these responses. This phenomenon is called endotoxin tolerance, but the mechanisms underlying it remain unclear. The subfornical organ (SFO) rapidly produces proinflammatory cytokines including interleukin-1 beta (IL-1 beta) in response to peripherally injected LPS, and repeated LPS injection attenuates IL-1 beta production in the SFO, indicating that the SFO is involved in endotoxin tolerance. The purpose of this study is to investigate features of the IL-1 beta source cells in the SFO of LPS-non-tolerant and LPS-tolerant mice. We first established the endotoxin-tolerant mouse model by injecting LPS into adult male mice (C57BL/6J). Immunohistochemistry was performed to characterize IL-1 beta-expressing cells, which were perivascular macrophages in the SFO. We depleted perivascular macrophages using clodronate liposomes to confirm the contribution of IL-1 beta production. To assess the effect of LPS pre-injection on perivascular macrophages, we transferred bone marrow-derived cells obtained from male mice (C57BL/6-Tg (CAG-EGFP)) to male recipient mice (C57BL/6N). Finally, we examined the effect of a second LPS injection on IL-1 beta expression in the SFO perivascular macrophages. We report that perivascular macrophages but not parenchymal microglia rapidly produced the proinflammatory cytokine IL-1 beta in response to LPS. We found that peripherally injected LPS localized in the SFO perivascular space. Depletion of macrophages by injection of clodronate liposomes attenuated LPS-induced IL-1 beta expression in the SFO. When tolerance developed to LPS-induced sickness behavior in mice, the SFO perivascular macrophages ceased producing IL-1 beta, although bone marrow-derived perivascular macrophages increased in number in the SFO and peripherally injected LPS reached the SFO perivascular space. The current data indicate that perivascular macrophages enable the SFO to produce IL-1 beta in response to circulating LPS and that its hyporesponsiveness may be the cause of endotoxin tolerance.