The effects of 3-methylcholanthrene (MCA), 5,6-benzoflavone (betaNF), 7,8-benzoflavone (alphaNF) and pregnenolone 16alpha-carbonitrile (PCN) upon aryl hydrocarbon hydroxylase (AHH) were determined in primary mammary gland epithelial cell cultures prepared from the C3Hf-/Ki mouse. MCA elevated AHH activity by 3--4 fold after 24 h of treatment; alphaNF produced a 50% inhibition. The specific activity of AHH in these cells was elevated by 6 h after exposure to MCA; enzyme activity was still maximally elevated after 48 h. The effects of MCA were also investigated in a group of mammary cell lines, one of which was derived from a control virgin mouse, the MCG V14; 3 of which arose from mammary tumors, MCG T10, MCG T14 and MCG T19; and 2 of which were sublines developed from hyperplastic alveolar nodules, HAN-1 and HAN-2. Induction was seen in all lines at 24 h, with the MCG T14 being the most responsive and the HAN-2, the least. Although the MCG T19 tumor cells did respond in culture, when implanted in the mouse, the AHH of the subsequent tumor was not elevated upon administration of MCA in vivo.