Halogenated dimethyl bipyrroles (HDBPs) recently have been identified as a group of environmentally persistent naturally produced bioaccumulative organohalogens. The ability of these compounds to activate the aryl hydrocarbon receptor (AhR) signaling pathway in vitro was examined. Induction of cytochrome P4501A (CYP1A), measured as ethoxyresorufin-O-deethylase (EROD) activity, and porphyrin accumulation were monitored after exposure of chick embryo hepatocytes to three individual HDBP congeners and two HDBP mixtures. All HDBP congeners and mixtures tested caused induction of EROD activity. Induction equivalency factors, calculated as the ratio of the effective concentration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to induce 10 and 50% of the maximal EROD induction to that of the HDBP congener or mixture ranged from 0.0004 to 0.05. Halogenated dimethyl bipyrroles also caused porphyrin accumulation, which increased with exposure time of the hepatocytes and molar bromine content of the HDBP congener or mixture. Heptacarboxylporphyrin III and corproporphyrinogen III were the most abundant porphyrins observed after an exposure period of 48 h. The individual HDBP congeners and mixtures also inhibited binding of [3H]TCDD to chicken and mouse AhR. Binding of [3H]TCDD to chicken AhR was inhibited approximately 20% by tetra- to hexabrominated congeners at concentrations of 20 to 40 microM; binding to mouse AhR was inhibited 40 to 50% by tri- and tetrabrominated dimethyl bipyrroles (DBPs) at 10 microM. The relative affinity of the hexabrominated DBP congener (vs TCDD) for binding to chicken AhR was estimated to be 0.000017. The results of this work illustrate that HDBPs act as agonist ligands for the AhR in the same manner as many anthropogenic organohalogens.