We identified cells of the developing superior cervical ganglia at postfertilization in living DbH transgenic fish and in whole mount in situ hybridization arrangements with dbh and th riboprobes, revealing that EGFP expression in the developing embryonic PSNS of this transgenic line recapitulates the normal endogenous expression patterns of dbh and th. By 80 hpf, EGFP was apparent within the superior cervical ganglia, in addition to in non PSNS dopaminergic neurons, including the medulla oblongata and cranial ganglia. By contrast, most MYCN transgenic embryos failed to convey Celecoxib price a detectable level of EGFP fused to human MYCN inside the superior cervical ganglia at 80 hpf, even though the fusion protein was clearly expressed in non PSNS tissues, and in most animals, the lack of detectable sympathoadrenal cells continued through 10 dpf. Having less EGFP expression is consistent with the substantially paid down variety of sympathoadrenal cells in MYCN embryos suggested by the loss of cells with endogenous th and dbh RNA expression by whole mount in situ hybridization. The absence of cells expressing EGFP MYCN under control of the dbh promoter can reflect either MYCN induced apoptosis or a charge in sympathoadrenal progenitor cell differentiation, since th and dbh Meristem are markers for differentiated sympathoadrenal cells. To differentiate between these possibilities, we first performed TUNEL and anti activated Caspase 3 staining on parts of 36, 51, and 72 hpf MYCN versus DbH transgenic fish. We found no proof TUNEL or anti activated Caspase 3 positive cells in the superior cervical ganglia or places where sympathoadrenal cells would be likely to form, suggesting that the absence of detectable sympathoadrenal cells is not due to cell death, but instead to a failure to start the PSNS developmental program as of this early time in development. To try this possibility, we performed entire mount in situ hybridization at 80 hpf and 54 hpf for term of the zash1a, phox2b, and AP 2 Everolimus 159351-69-6 alpha genes, which encode transcription factors needed for sympathoadrenal cell specification and maintenance. Each of these sympathoadrenal cell progenitor guns was readily detectable in the superior cervical ganglia location of control embryos, but undetected in MYCN transgenic embryos at these stages, indicating that specification of the earliest recognizable sympathoadrenal cell progenitors was blocked by expression of the EGFP MYCN fusion gene. The reduction of sympathoadrenal cell development by EGFP MYCN seems to be tissue specific, since expression of the EGFP MYCN by non PSNS dopaminergic neuronal cells in these embryos was largely unaffected, including expression by cells of the medulla oblongata, locus coeruleus, and cranial ganglia.