We have shown that bronchoalveolar epithelial A₁-adenosine receptors (A₁-AdoR) are activated in influenza A virus-infected mice. Alveolar macrophages and neutrophils also express A₁-AdoRs and we hypothesized that activation of A₁-AdoRs on these cells will promote macrophage and neutrophil chemotaxis and activation and thereby play a role in the pathogenesis of influenza virus-induced acute lung injury. Wild-type (WT) C57BL/6 mice, congenic A₁-AdoR-knockout (A₁-KO) mice, and mice that had undergone reciprocal bone marrow transfer were inoculated intranasally with 10,000 PFU/mouse influenza A/WSN/33 (H1N1). Alternatively, WT mice underwent daily treatment with the A₁-AdoR antagonist DPCPX from one day prior to inoculation. Infection increased BALF adenosine comparably in WT and A₁-KO mice. Infection of WT mice resulted in reduced carotid arterial O₂ saturation (hypoxemia), lung pathology, pulmonary edema, reduced lung compliance, increased basal airway resistance, and hyperresponsiveness to methacholine. These effects were absent or significantly attenuated in A₁-KO mice. BALF leukocytes, IFN-γ, and IL-10 were significantly reduced in infected A₁-KO mice, but KC, IP-10, and MCP-1 were increased. Reciprocal bone marrow transfer resulted in WT-like lung injury severity, but BALF leukocytes only increased in WT to A₁-KO mice. Hypoxemia, pulmonary edema, and BALF alveolar macrophages, neutrophils, IFN-γ, and IL-10 were reduced in DPCPX-treated WT mice. Viral replication did not differ between mouse strains or treatment groups. These findings indicate that adenosine activation of leukocyte A₁-AdoRs plays a significant role in their recruitment to the infected lung and contributes to influenza pathogenesis. A₁-AdoR inhibitor therapy may therefore be beneficial in patients with influenza-induced lung injury.