During Gram-negative endotoxemia, precise regulation of monocyte/macrophage (Mφ) responsiveness to lipopolyssaccharide (LPS) is critical to preserve host defense while avoiding complications such as organ failure and death. We will discuss regulation of LPS-Mφ interactions by LPS-binding plasma proteins and by LPS-induced changes in Mφ responsiveness. Upon exposure to plasma, LPS binds to either lipoproteins or LPS-binding protein (LBP; a 60-kilodalton glycoprotein with a high-affinity binding site for the lipid A moiety of rough and smooth LPS). The LPS-LBP complex stimulates the Mφ by binding to its cellular receptor, CD 14 (a monocyte/Mφ-specific, phos-phatidylinositol-anchored surface glycoprotein). Pretreatment of whole blood with anti-CD 14 monoclonal antibody reduces the responsiveness of monocytes to LPS [determined by tumor necrosis factor-α (TNF-α) release] at least 10-fold. Similarly, cellular responsiveness to LPS is diminished at least 100-fold by depletion of plasma LBP with anti-LBP antibody. Compared to LPS-LBP induction of TNF-α, LPS-lipoprotein complexes are as much as 10,000-fold less active. Thus, partitioning of LPS between LBP and lipoproteins markedly influences Mφ responsiveness to LPS. LPS also directly induces Mφ hyporesponsiveness to itself by a process known as adaptation; exposure of Mφ to < 100 pg LPS/ml (subthreshold for TNF induction) for 6–9 reduces the sensitivity of the Mφ to subsequent challenge up to 1,000-fold, so that 1 µg/ml rather than 1 ng/ml of LPS is required for maximal induction of TNF-α. LPS-adapted Mφ are specifically unresponsive to LPS (rough or smooth), yet remain fully responsive to heat-killed Staphylococcus aureus. The LPS-induced increase in steady state levels of TNF mRNA characteristic of control Mφ is not observed in adapted Mφ. This is not due to increased mRNA degradation or to autocrine inhibiton of Mφ function by secretory products. Exposure of LPS-adapted Mφ to LBP increases the sensitivity of these cells to LPS. Thus, multiple pathways regulate host responsiveness to LPS and establish the balance between defense and injury.