The effects of the murine lymphomagen, 1,3-butadiene (BD), on the proliferation and differentiation of hematopoietic stem cells were examined in male B6C3F1 mice. Exposure to 1250 ppm BD for 6 weeks resulted in no demonstrable alteration in the frequency of spleen colony-forming units (CFU-S); however, colonies derived from treated animals were smaller than those from controls. The absence of any difference in the frequency of CFU-GM after 6 weeks exposure suggests that BD produces an alteration in the relative proportion of immature to mature pluripotent stem cells in BD-exposed animals. This was confirmed by the examination of the effects of BD on stem cell development in long-term bone marrow culture. After 14 days, the number of CFU-GM derived from cultures of animals exposed for 6 weeks was reduced compared to controls. However, at 28 days an increase relative to controls was observed. This shift in the course of differentiation of the granulocyte/macrophage precursor cell, as assessed by the CFU-GM, provides further evidence that there is an increase in the relative frequency of primitive or immature stem cells in BD-treated mice. After a 30-31 week exposure to BD, a decrease in the numbers of both CFU-S and CFU-GM was observed. These findings indicate that BD causes alterations in stem cell development and suggest that alterations in bone marrow stem cells may play an essential role in the pathogenesis of BD-induced thymic lymphoma.