TCDD-induced transcription and developmental toxicity in zebrafish

Developmental exposure of zebrafish to 2,3,7,8-tetmchlorodibenzo- p-dioxin (TCDD) causes edema, anemia, hemorrhage and ischemia associated with arrested growth and development. Heart and vasculature development and function are severely impaired and jaw malformations occur secondary to inhibited chondrogenesis. This profile of developmental toxic responses is observed in the larval form of all freshwater fish species exposed to TCDD at the embryonic stage of development. Components of the aryl hydrocarbon receptor/aryl hydrocarbon receptor nuclear translocator (AHR/ARNT) signaling pathway that mediate TCDD toxicity have been identified and functionally characterized in zebrafish. The zebrafish has two forms of the AHR, zfAHR1 and zfAHR2. To determine whether one or both forms mediate toxicity, an antisense morpholino approach was used to knockdown translation of zfAHR2. The zfahr2 morpholino protected embryos from TCDD-induced developmental toxicity, demonstrating that zfAHR2 mediates TCDD developmental toxicity in zebrafish. AHR is a ligand-activated transcription factor, and the AHR/ARNT dimer likely mediates TCDD toxicity by binding specific DNA response elements in promoters of target genes resulting in altered expression. To determine whether cytochrome P4501A (CYP1A), the most well characterized AHR/ARNT target gene, mediates TCDD developmental toxicity in zebrafish, an antisense morpholino approach was also used to knockdown translation of zfCYP1A. Unlike the zfahr2 morpholino, the zfcyp1a morpholino did not protect against TCDD-induced developmental toxicity in zebrafish even though it prevented zfCYP1A upregulation more effectively than the zfahr2 morpholino. This suggests other TCDD-activated AHR/ARNT target genes are responsible for mediating TCDD developmental toxicity in zebrafish. Microarray analysis of global gene expression changes induced by TCDD in the hearts of zebrafish larvae was performed to identify other TCDD-activated AHR/ARNT target genes that mediate TCDD-induced cardiovascular toxicity. TCDD exposure quickly elicited a transcriptional response in the hearts of exposed zebrafish larvae that preceded a TCDD-induced decrease in cardiac output and cardiac myocyte cell number. This response included expression changes in genes that regulate myocyte differentiation, contractile function, and proliferation. These TCDD responsive genes are strong putative AHR/ARNT gene targets that may mediate TCDD-induced cardiovascular toxicity in zebrafish larvae.