Autophagy, a homeostatic mechanism, is crucial in maintaining normal cellular function. Although dysregulation of autophagic processes is recognized in certain diseases, it is unknown how maintenance of cellular homeostasis might be affected by the kinetics of autophagic activity in response to various stimuli. In this study, we assessed those kinetics in lung adenocarcinoma (A549) cells in response to exposure to nanoparticles (NP) and/or Rapamycin. Since NP are known to induce autophagy, we wished to determine if this phenomenon could be a driver of the harmful effects seen in lung tissues exposed to air pollution. A549 cells were loaded with a fluorescent marker (DAPRed) that labels autophagosomes and autolysosomes. Autophagic activity was assessed based on the fluorescence intensity of DAPRed measured over the entire cell volume of live single cells using confocal laser scanning microscopy (CLSM). Autophagic activity over time was determined during exposure of A549 cells to single agents (50 nM Rapamycin; 80 µg/mL, 20 nm carboxylated polystyrene NP (PNP); or, 1 µg/mL ambient ultrafine particles (UFP) (<180 nm)), or double agents (Rapamycin + PNP or Rapamycin + UFP; concomitant and sequential), known to stimulate autophagy. Autophagic activity increased in all experimental modalities, including both single agent and double agent exposures, and reached a steady state in all cases ~2 times control from ~8 to 24 hrs, suggesting the presence of an upper limit to autophagic capacity. These results are consistent with the hypothesis that environmental stressors might exert their harmful effects, at least in part, by limiting available autophagic response to additional stimulation, thereby making nanoparticle-exposed cells more susceptible to secondary injury due to autophagic overload.