Therefore, the observed increase in HRinduced apoptosis in HSF1 knockdown embryos may be in a cell population that normally contains higher levels of HSF1 than their fully differentiated counterparts. However, brains of embryos at the stage of development used in our experiments express markers for differentiated neurons . In addition, retinal cell differentiation is evident in the zebrafish embryo by 48 hpf . Therefore, our results demonstrating an essential role for HSF1 in cell survival after IR are CP-690550 likely relevant to differentiated brain and eye tissues. Rapamycin expression of heat shock proteins, such as Hsp27 and Hsp70, has been shown to protect cells against IR in a variety of model systems. However, results of the present study show that both Hsp70 and Hsp27 are expressed in HSF1 knockdown embryos, yet these embryos none-the-less display elevated levels of apoptotic cell death after HR compared to control MO injected animals. These data suggest that there are essential functions for HSF1 other than the promotion of heat shock protein expression in the brain and eye, and are consistent with recent studies showing increasingly complex roles for HSF1 in development and tissue homeostasis. For example, Drosophila HSF is capable of binding to nearly 3% of genes and recent reports indicate that vertebrate HSF1 also binds to and activates numerous genes outside the canonical stress response pathway . Additionally, in HSF1-null mice, cardiac and renal cells redox homeostasis is impaired through still poorly defined mechanisms. Inability to properly regulate redox states could make highly metabolically active tissues, such as found in the brain, unusually vulnerable to the oxidative damage of IR. HSF1 null mice also have altered glucose uptake and metabolism . Animals lacking HSF1 may be unable to undergo critical alterations in glucose metabolism during hypoxic challenge. Therefore, it is unsurprising that the lack of HSF1 has deleterious effects in zebrafish embryos despite the presence of Hsp27 and Hsp70. Hsp27 and Hsp70 expression may be induced by IR through a mechanism other than HSF1 activation Our results are consistent with previous studies showing the ability of HSF1 to protect cells against IR, but are novel in showing that Hsp70 and Hsp27 expression persists after HR despite substantial reduction in HSF1 expression. It is possible that these results reflect limitations of our methods. For example, although we demonstrate effective knockdown of HSF1 by Western blot, morpholino microinjection, unlike gene deletion methods, does not completely eliminate HSF1 expression. The quantity of morpholino injected into embryos in the present study was sufficient to inhibit heat shock-induced upregulation of Hsp27 .