In Ovo induction of hepatic ethoxyresorufin o-deethylase activity by 2,3,7,8-tetrachlorodibenzo-P-dioxin in four avian species
Creators: name: Sanderson, John Thomas
Publisher: name: University of British Columbia
Year: 2009

description: The Canadian Wildlife Service monitors levels of polychiorinated aromatic hydrocarbons in great blue heron (Ardea herodias) and double-crested cormorant (Phalacrocorax auritus) eggs in British Columbia (B.C.) as biological indicators of environmental contamination. This thesis assessed the spatial and temporal effects of environmental contamination with polychiorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs) and biphenyls (PCBs) on hepatic microsomal ethoxyresorufin 0-deethylase (EROD) activities and other biological parameters in heron hatchlings from colonies in B.C and cormorant hatchlings from colonies across Canada. Also, the induction of hepatic EROD activities by environmental levels of polychiorinated aromatic hydrocarbons were compared to in ovo dose-response studies, using 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), in the laboratory. Finally, the differences in EROD-inducing potency of TCDD among avian species were compared with the affinities of TCDD for the avian hepatic Ah receptors. In the herons, levels of PCDDs and PCDFs had decreased considerably since 1988, particularly in the Crofton colony. Concomitantly, there was a decrease in hepatic EROD activity and incidence of chick edema, an increase in body weight and organ weights, and improvement of the reproductive success of the Crofton colony. The reduction in severity of the effects observed in the contaminated colonies in the recent collections, accompanied by the decline in levels of PCDDs and PCDFs, was consistent with the dose-response relationships determined in 1988. Regression analysis demonstrated a significant positive relationship between hepatic EROD activity (pmole/minlmg protein) and TCDD level =(2O.r49; p&lt;O.00005; n=54). In the cormorants, hepatic EROD activity was positively related to the sum of TCDD-toxic equivalents =(2O.r69; p&lt;O.00005; n25). Immunoblots showed crossreactivity of monospecific antibodies raised against rat cytochrome P-45 0 1 Al (CYP 1 Al) with a hepatic microsomal protein in the heron and cormorant hatchlings. Staining of the band increased with increased EROD activity and increased exposure of the birds. These results support the use of avian hepatic microsomal EROD activity as an index of CYP1A1 induction by environmental levels of polychiorinated aromatic hydrocarbons, and as a useful biomarker of the extent of exposure to such chemicals. Furthermore, the induction of CYP 1A1 observed in the heron and cormorant indicates that the Ah receptor-mediated process, by which TCDD and related chemicals exert many of their toxicities, has been activated. The in ovo hepatic EROD-inducing potency of TCDD, as an indicator of sensitivity to Ah receptor-mediated toxicities, was determined in the domestic chicken (Galius gallus), domestic pigeon (Columba livia), great blue heron and double-crested cormorant. Doseresponse curves were produced by injecting various doses of 3H-TCDD (in corn oil) into the air sac of developing eggs, during the latter third part of incubation. After hatching, yolk, blood and a portion of liver were analysed for 3H-TCDD, using liquid scintillation counting. There were no major differences in organ distribution of TCDD among hatchlings of the four avian species. The ED50 for EROD induction by TCDD was about 1 ‘/2 order of magnitude lower in the domestic chick (0.1 pg/kg), a species known to be sensitive to TCDD toxicity, than in the heron and cormorant (3-10 pg/kg). The pigeon was similar to the wild bird species. Consistent with this, receptor binding studies showed that the affinity of 3H-TCDD for the hepatic Ah receptor was about 10 to 15 times higher in the chick (Kd=O. 7S and 1.6 nM) than in the other avian species (pigeon: Kd=l 1 and 14 nM; heron: Kd=lO and 20 nM; cormorant: Kd=l 2 and 16 nM). Receptor binding affinities in pigeon, heron and cormorant were similar to that reported in human placenta. Subcutaneous edema was observed in TCDD-treated hatchlings of the chick, heron and cormorant, but not in the pigeon hatchling, at the dose-range examined. Comparison of the laboratory dose-response curves with environmental exposures to TCDD and related chemicals indicates that hepatic EROD induction is at the lower end of the linear part of the dose-response curves in both the great blue heron and double-crested cormorant. A further increase in levels of TCDD and similarly acting compounds would lead to a large increase in hepatic EROD activity and further increases in other Ah receptormediated toxicities, such as body weight loss and edema. Key Words: Avian, Ah receptor, biological monitoring, dose-response, double-crested cormorant, ethoxyresorufin 0-deethylase, great blue heron, 2,3,7, 8-tetrachlorodibenzo-pdioxin, toxic equivalents.