Study of hepatic cytochrome P450 system in Richardson ground squirrels
Richardson ground squirrels (gophers) are pests on the prairies that cause considerable agricultural and ecological damage. Traditional control methods such as the rodenticides strychnine, zinc phosphide, and anticoagulants, have proven ineffective in reducing gopher densities. In additional, current gopher control methods have the significant potential to cause primary and secondary toxicity to non-target animals. Thus, alternative methods for toxicological control of gophers are needed to mitigate these concerns. Present studies focused on the cytochrome P450 (CYP450) enzyme system responsible for xenobiotic detoxification in gophers. In vitro hepatic microsomal systems and HPLC analysis were used to elucidate general metabolic characteristics of major gopher xenobiotic metabolizing pathways. We found that the content and activity of individual components of the CYP450 system including CYP450, cytochrome b5, and NADPH-cytochrome P450 reductase in liver microsomal preparations were higher in gophers exposed to toxins used to control their population than in naïve (unexposed) gophers. When in vitro CYP450 mediated activities for five substrates [coumarin and aniline aromatic hydroxylation, 7-methoxycouamrin O-demethylation, and N-methylaniline, and N,N-dimethylaniline N-demethylation] were measured, naïve gophers were identified to have higher specific activity but similar whole body activity compared to the exposed gophers. Furthermore, there was a clearly identifiable sub-population of “poor metabolizers” showing considerably lower CYP450 activity within the gopher samples studied. Clotrimazole was found to be a potent inhibitor of several substrates of CYP450 enzyme-mediated reactions, which included aniline aromatic hydroxylation, N-methylaniline and N,N-dimethylaniline N-demethylation, and 7-methoxycoumarin O-demethylation. The cytotoxicity of above compounds was tested using freshly isolated gopher hepatocytes. The results showed that each compound caused considerable cytotoxicity to gopher hepatocytes. Addition of clotrimazole to the freshly isolated hepatocyte suspension increased the cytotoxicity of all tested compounds. In conclusion, gophers may develop resistance to current chemical control methods through the enhancement of CYP450 system content, which can compensate the loss of enzyme activity. Furthermore, clotrimazole is a potent cytochrome P450 inhibitor, which increases the cytotoxicity caused by given compounds in gopher livers. The concept of using CYP450 enzyme inhibitor in combination with another chemical whose elimination depends on CYP450 metabolism to improve current gopher control method has practical importance.
Cytochrome P450, Richardson ground squirrels
Master of Science (M.Sc.)
Animal and Poultry Science
Animal and Poultry Science