Lipopolysaccharide (LPS) tolerance is a state of refractoriness towards a second stimulation by LPS after a preceding stimulation. LPS is recognized by Toll‐like receptor‐4 (TLR‐4), which belongs to a group of pattern recognition receptors mediating activation of innate immunity by microbial components. To date, it is not known in detail to what extent other TLR‐dependent stimuli also induce tolerance and whether preceding and challenging stimuli are interchangeable. We have examined tolerance induction in detail for lipoteichoic acid (LTA), LPS and CpG‐DNA, which are recognized by TLR‐2, ‐4 and ‐9, respectively. In RAW264·7 macrophages, all three stimuli induced tolerance towards a subsequent challenge with the same stimulus used for priming, as well as cross‐tolerance towards subsequent challenge with other stimuli signalling via different TLRs. However, whereas LPS/LTA cross‐tolerance was also functional in an in vivo model of galactosamine (GalN)‐primed liver damage, pretreatment with CpG only protected against GalN/CpG challenge and failed to induce cross‐tolerance for LPS and LTA. CpG‐DNA pretreatment even enhanced tumour necrosis factor (TNF)‐α production and liver damage upon subsequent challenge with LPS or LTA. Stimulation with CpG‐DNA resulted in a peculiar sensitization for interferon (IFN)‐γ secretion. The data indicate that, in contrast to in vitro macrophage desensitization, the in vivo consequences of repeated TLR stimulation greatly differ amongst different TLR ligands.