Tumor necrosis factor (TNF)–α, interleukin (IL)–1β, and nitric oxide (NO) may play a role in lipopolysaccharide (LPS)–induced cardiac depression. Toll-like receptor–4 (TLR-4) mediates the cytokine response to LPS in immune cells. TLR-4 also is expressed in human and murine myocardial tissue. Therefore, the hypothesis that LPS induces proinflammatory cytokines in the heart via TLR-4 was tested. C3H/HeJ (TLR-4 deficient) and C3HeB/FeJ mice were studied. LPS induced a robust increase in myocardial TNF-α and IL-1β mRNA in C3HeB/FeJ mice. The response in C3H/HeJ mice was blunted and delayed. Myocardial TNF-α and IL-1β protein levels were higher in C3HeB/FeJ mice, as were inducible NO synthase protein and NO production. Activation of myocardial NF-κB was observed within 30 min in C3HeB/FeJ mice but not in C3H/HeJ mice. These findings suggest that myocardial TLR-4 is involved in signaling cytokine production within the heart during endotoxic shock