Unmethylated CpG motifs in bacterial DNA or synthetic oligodeoxynucleotides (ODN) potently stimulate the innate immune system, and they are recognized by Toll-like receptor 9 (TLR9), which is expressed by monocytes/macrophages, dendritic cells, and B cells. However, it is unknown whether alveolar macrophages (AMs) express functional TLR9. To clarify this, we analyzed mRNA expressions of TLRs in murine AMs by real-time polymerase chain reaction, and compared with those in other tissue macrophages and lung antigen-presenting cells. In addition, we determined the sensitivity of these cell populations to CpG-ODN. Interestingly, TLR9 mRNA was almost absent in AMs, but highly expressed in bone marrow-derived macrophages and peritoneal macrophages, whereas TLR2 and TLR4 were present in all macrophage populations. Consistent with the receptor expression, AMs showed no sensitivity to CpG-ODN, whereas other macrophage populations secreted tumor necrosis factor alpha, interleukin 12 p40, and interleukin 6, and enhanced expression of CD40, CD80, and CD86, in response to CpG-ODN. Lung dendritic cells and B cells highly expressed TLR9 mRNA and responded to CpG-ODN. These results indicate selective loss of TLR9 expression in AMs with no sensitivity to CpG-ODN, suggesting that dendritic cells and B cells play a role in the immune response against bacterial DNA in the lung.