Previous studies have shown that exposure of pathogen-free C57BL/6N mice to 5 or 10 ppm NO2 increases the severity of murine respiratory mycoplasmosis and that this effect is associated with decreased intrapulmonary killing of Mycoplasma pulmonis. The purposes of the present studies were to determine the effects of doses of NO2 lower than 5 ppm on pulmonary clearance and to provide experimental links between NO2 exposure, defects in intrapulmonary killing, and alterations in alveolar macrophages. Exposure to less than 5 ppm NO2 had no effect on intrapulmonary killing of M. pulmonis. Bronchoalveolar lavage cells killed M. pulmonis in vitro only if they were allowed to associate with mycoplasmas in vivo. Prior exposure to NO2 abrogated killing in this in vivo-in vitro model. More than 95% of the BAL cells were macrophages, and more than 98% of the cell-associated mycoplasmas were on or in alveolar macrophages. Immediately after exposure, the viability of alveolar macrophages was 89 +/- 4% in the control group, 56 +/- 19% in the group receiving M. pulmonis alone, 23 +/- 7% in the group receiving 10 ppm NO2, and 16 +/- 6% in the group receiving both M. pulmonis and NO2 exposures. Viability was significantly decreased following exposure to 10 and 5 ppm NO2 but not following exposure to 2 ppm. Both viability and intrapulmonary killing were depressed at 3 days after exposure to NO2 but were normal by 7 days after exposure. The cellular target of NO2 exposure in relation to intrapulmonary killing of M. pulmonis appears to be the alveolar macrophages.