The herbicide 3,4-dichloropropionanilide (propanil) has several well-documented neurotoxic and immunotoxic effects on mice. We report here a detailed characterization of the effects of propanil exposure on the thymus. We found that at doses of 100-200 mg/kg, propanil induces significant thymic atrophy between 2 and 7 days postexposure. This atrophy is characterized by a decrease in thymus/body ratio and a decrease in cellularity. Flow cytometric analyses of thymuses from propanil- and vehicle-treated mice indicate that the CD4(+) CD8(+) population of immature cells, is most significantly decreased in propanil-exposed mice. We performed cell cycle analysis of thymocyte populations using two-color surface staining and the DNA binding dye 7-aminoactinomycin D to determine whether thymic atrophy was associated with changes in the percentages of cells in the S, G2, and M phases of the cell cycle. We found a high percentage of proliferating CD4(+)CD8(+) thymocytes 4 days after exposure. Thus, recovery of the thymus occurs following increases in thymocyte proliferation, most notably the immature CD4(+) CD8(+) thymocytes. We tested the hypothesis that glucocorticoids play a role in the observed atrophy by examining thymuses in adrenalectomized, propanil-treated mice. No atrophy was observed in those animals. These results suggest that propanil has an immunotoxic effect on the thymus that appears to be mediated, in part, by endogenous glucocorticoids.