Submitted to: Food and Chemical Toxicology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 29, 2005
Publication Date: May 1, 2006
Citation: Roberge, M.T., Hakk, H., Larsen, G.L. 2006. Cytosolic and localized inhibition of phosphodiesterase by atrazine in swine tissue homogenates. Food and Chemical Toxicology 44:885-890. Interpretive Summary: Atrazine is one of the most widely used herbicides in the United States. Claims have been made that Atrazine disrupts the proper maturation of animals, most notably amphibians (frogs) and rats. It was speculated that Atrazine disrupts the endocrine system, a system of organs that control hormonal signals during adolescence and adulthood. However, we showed previously that endocrine disruption did not occur directly with the estradiol/estrogen receptor system, but that Atrazine inhibited phosphodiesterase, an enzyme that controls signals within cells. To further our understanding on how this happens we collected brain, liver, lung, kidney, and heart tissues from three different pigs at slaughter. The tissues were ground up in a special solution to keep the phosphodiesterase enzymes operational and their activity was observed in the presence of known inhibitors and Atrazine. We found that phosphodiesterase in heart, lung and brain could be inhibited by Atrazine but not kidney or liver. In addition, we separated the cell membranes and organelles of the cells from the cytosol (liquid inside), only the cytosol from the heart was inhibited by Atrazine. There are more than 50 different types of phosphodiesterases that are preferentially distributed among tissues. These experiments tell us that Atrazine may inhibit specific types of phosphodiesterases present in specific tissues. This information is important because it lays the groundwork to look for chemical markers in live animals in order to determine if Atrazine really is an endocrine disruptor.
Technical Abstract: Atrazine (ATR) significantly inhibited phosphodiesterase (PDE) in crude homogenates of swine heart, brain, and lung but not liver or kidney tissues. Except for heart, PDE activities in the cytosolic fraction of the tissue homogenates were not affected by ATR. The inhibition of the PDE activity in the cytosol from heart homogenate was not significantly different between ATR and a non-specific PDE inhibitor, 3-isobutyl-1-methylxanthine (IBMX). Dixon plots of the crude tissue homogenates showed that heart and brain were inhibited via two different mechanisms (competitive or mixed inhibition, and noncompetitive inhibition respectively), suggesting that ATR may be a semi-specific PDE inhibitor. Furthermore, in crude tissue homogenates, ATR did not inhibit PDE as effectively as IBMX suggesting that there are ATR-susceptible and ATR-nonsusceptible forms of PDE. Association constants for ATR were 65 µM for heart and 325 µM for brain. The stability of the activity of PDE was affected by freezing, requiring the use of only freshly prepared tissue homogenates.