The anti-HIV agents AZT (zidovudine) and ddl (dideoxyinosine) are being used clinically during pregnancy. The toxicity of these agents to the fetus and placenta remains a concern because few human pregnancy exposure data are available, and pregnant rodent studies with AZT indicate increased embryonic resorptions and developmental arrest. The current study used a human choriocarcinoma cell line (JAr), which exhibits many characteristics of the early placenta, to assess the effects of a single 24 h exposure of 7.6 or 0.076 mM AZT, and the effects of a single 24 h exposure of 7.6 or 0.076 mM ddI upon cell proliferation and hormone production of human chorionic gonadotropin (hCG), estradiol (E2), and progesterone (P4). The higher concentration of AZT and ddI produced significant (P < 0.025) reductions in cell numbers and growth rate while producing significant increases in hormone production (hCG, E2, and P4). The lower concentration of AZT and ddI produced significant increases in E2 production, but no changes in cell numbers, hCG, or P4. Because placental cells require androgen precursor for E2 synthesis, exogenous androstenedione was added to confirm observations of increased estradiol synthesis after AZT or ddl exposure. These results demonstrate that single 24 h high dose exposures of AZT or ddI produce significant inhibition of cell proliferation and alterations in hormone production in this paradigm of human placental cells.