Behrents Hartney, Kristine, 105 Bematchez, Louis, 247 Breitburg, Denise L., 97 Brown, Larry R., 223 Carl, Leon M., 239 Christensen, V., 87 Cooper, Scott D., 285 Copp, Gordon H., 1 Dodson, Julian J., 247 Donaldson, Terry J., 295 Dudley, Tom L., 285 Dunson, William A., 303 Farr, James A., 39,119 Fox, Michael G., 129 Gonzalez, Richard J., 303 Gunnarsson, Karl, 159 Gutherz, Elmer J., 277 Hales, L. Stanton, Jr., 153 J&vi, Torbjiim, 149 Jones, Robert S., 277 Keast, J. Allen, 129 Kemp, Alain, 247 Kiorboe , T., 87 Kraft, Melvin, 239 Lammens, Eddy H.R.R., 63 Matlock, Gary C., 277 McManus, Michael, 119 Mikhail, Mansour Y., 49 Munk, Peter, 87 Nelson, Walter R., 277 Ochi, Haruki, 257 Pankhurst , Neville W., 201 Parrish, Julia K., 79 Petersen, Christopher W., 213 Power, Mary E., 285 Quinn, Thomas P., 311 Rhude, Larry, 239 Ruggerone, Gregory T., 143 Sale, Peter F., 77 Sigurj&sd&tir, Hrefna, 159 Smith, C. Lavett, 237 Smith, Marcus P.L., 29 Swainson, Robert J., 129 Travis, Joseph, 39, 119 Trexler, Joel C., 119 Visser, Jentsje T., 63 Welch, Harold E., 49 Winemiller, Kirk O., 177

Previous studies show that changes in estrogen (ER) and glucocorticoid receptor (GR) function in rainbow trout (Oncorhynchus mykiss) oocytes modulate the growth performance phenotype of embryo and juvenile progeny; the present study was undertaken to determine whether this altered growth performance is associated with changes in the expression of several growth-related genes in early-stage embryos. Unfertilized oocytes were incubated in the presence of various combinations of GR and ER agonists and antagonists; the oocytes were then fertilized and the expression of genes that encode for six nuclear receptor superfamily (NRS) proteins (GR1, GR2, ERα, ERβ, TRα, and TRβ) and the two IGF peptides (IGF1 and IGF2) were measured in the 7-, 13-, and 26-dpf embryos. By day 26 of embryogenesis, the expression of the six NRS-related genes of interest and that of igf2 were significantly enhanced in embryos reared from ER agonist- or ER antagonist-treated oocytes, regardless of whether the GR agonist, cortisol, was also included in the initial oocyte incubation medium. Conversely, the igf1 expression pattern among treatment groups was significantly enhanced in the cortisol-only treatment group and in the ER antagonist and GR antagonist groups that were co-incubated with cortisol. Additionally, in the ER agonist treatment groups igf1 expression was significantly inhibited when cortisol was included in the oocyte incubation medium. The findings show that a single in ovo exposure to the receptor agonists/antagonists markedly changed the programming of the expression of NRS-related and IGF-related genes of the early-stage trout embryos.

Epigenetics is the study of changes in gene expression patterns that occur without any modification of the underlying nucleotide sequence of the DNA. Modifications of the so-called epigenome include complex transient or permanent chemical changes of the DNA or histone proteins resulting in the suppression or enhancement of gene expression, together with an array of post-translational events that modify the translational products. Epigenomic programming (EP) of the genome is an essential component of embryonic development in animals from the totipotent fertilized egg to the pluripotent stem cells, stem cell differentiation and final tissue and organ formation. Many of these EP processes are influenced transiently and some permanently by environmental influences. In eutherian mammals, environmentally related EP of embryos is linked to permanent changes in the phenotype of the progeny, some of which have been associated with adult onset metabolic disorders. Moreover, because some of the epigenetic remodelling occurs in both the soma and germ line, the resultant phenotypic characteristics (some of which are linked to disease states) may be heritable. Although far less is known about the effects of environmentally linked EP on the ontogeny of fishes, the available information suggests that the EP processes are similar amongst all vertebrates, and there are clear parallels between fish and mammals that are discussed in this paper. Our perspective takes the well-established findings in mammals and uses them to proactively extrapolate to the as yet under-recognized implications of EP for fish biology and for fish production in intensive aquaculture.

Increased in ovo cortisol content of rainbow trout oocytes from ~3·5 to ~5·0 ng.oocyte(-1) before fertilization enhances the growth of embryos and juveniles and changes the long-term expression pattern of IGF-related genes. This study used embryos reared from oocytes enriched with cortisol and the glucocorticoid receptor (GR) antagonist, RU486, to determine whether the growth-promoting actions of cortisol involve GR protein activation and modulation of gr expression. Whole-mount in situ immunohistofluorescence studies of zygotes showed that enhanced oocyte cortisol increased the immunofluorescent GR signal and activated the relocation of GR from a general distribution throughout the cytoplasm to an accumulation in the peri-nuclear cytoplasm. In ovo cortisol treatment increased the number of embryonic cells within 48-h post-fertilization, and RU486 partially suppressed this cortisol stimulation of cell duplication. In addition, there was complex interplay between the expression of gr and igf system-related genes spatiotemporally in the different treatment groups, suggesting a role for GR in the regulation of the expression of development. Taken together, these findings indicate an essential role for GR in the regulation of epigenomic events in very early embryos that promoted the long-term growth effects of the embryos and juvenile fish. Moreover, the pretreatment of the oocyte with RU486 had a significant suppressive effect on the maternal mRNA transcript number of gr and igf system-related genes in oocytes and very early stage embryos, suggesting an action of antagonist on the stability of the maternal transcriptome.

Mid-vitellogenic stage rainbow trout (Oncorhynchus mykiss) ovarian follicles (both intact and yolk free (YF)), ovulated oocytes and embryos were co-incubated with [2,4,6,7-(3)H]cortisol for 18 h to determine the degree and nature of the metabolism and biotransformation of the glucocorticoid. There was evidence of the conversion of cortisol to the less biologically potent glucocorticoid, cortisone, and the formation of glucocorticoid sulphates (both cortisol and cortisone) for all cell and tissue samples, suggesting the presence of 11β-hydroxysteroid dehydrogenase (11β-HSD) and glucocorticoid sulphotransferase (GST) activity at all stages; however, GST activity was particularly marked in both intact and YF ovarian follicles, suggesting an important role of follicles in limiting the exposure of oocyte to maternal cortisol. As there was no evidence of 11β-HSD or GST activity in ovarian fluid, the findings affirm that ovarian follicles (probably the thecal and granulosa cells) provide a barrier against the transfer of cortisol to the oocytes by forming sulphated steroids, whereas ovulated oocytes and early embryos have a more limited capacity to either metabolize or conjugate cortisol and are therefore more vulnerable at the post-ovulatory and early embryonic stages to increases in exposure to the glucocorticoid.

Cryptobia salmositica is a pathogenic haemoflagellate of Pacific salmon, Oncorhynchus spp., on the west coast of North America. The in vitro multiplication of the parasite was significantly enhanced by the addition of cortisol (within a range consistent with physiological levels in salmonid fishes; 10-50 ng ml(-1)) to the culture medium (MEM supplemented with FBS). However, higher cortisol concentrations (100 and 200 ng ml(-1)) either had no enhancing effects or resulted in lower replication rates compared with the controls. The synthetic glucocorticoid, dexamethasone (Dex), also stimulated the replication of the parasite and mifepristone (RU486), a synthetic steroid that has glucocorticoid receptor (GR) antagonist properties, inhibited the stimulatory actions of both cortisol and Dex, when added to the medium at a concentration of 100 ng ml(-1) co-culture with cortisol or Dex. Furthermore, the dose-dependent effects of glucocorticoids (cortisol and Dex) on the multiplication of the haemoflagellate were correlated with the initial size of the inocula. The study revealed a novel relationship between the parasite and its host, in which the host's cortisol is used by the parasite to enhance its replication. Also, since C. salmositica responds to both native and synthetic glucocorticoids and to the GR antagonist, RU486, and exhibits a biphasic (hormetic) response to the amount of cortisol in the medium, we propose that the glucocorticoid exerts its effects via an interaction with GR-like proteins in C. salmositica that are functionally similar to those present in vertebrate cells.

Event Abstract Back to Event In ovo elevated oocyte cortisol content induces innate immune responses in rainbow trout (Oncorhynchus mykiss) early embryonic cells Mao Li1*, Spencer Russell2, John S. Lumsden2 and John F. Leatherland1 1 University of Guelph, Biomedical Sciences, Canada 2 University of Guelph, Pathobiology, Canada In most fish embryos, the functional acquired immunity begins after hatch (1), and the protection of the embryos relies on the innate immunity of early embryo cells (1, 2). The immune suppressive effects of cortisol are well established in the developed organism, but the effects of maternal cortisol on early embryo immunity, and the ontogeny of innate immune components are not well established (2, 3). Lectins and TLRs were studied because they are widely distributed among fish tissues, and responsive to pathogens (5, 6). Naturally-ovulated rainbow trout oocytes were incubated in cortisol-enriched ovarian fluid for 3-h to examine the effects of the cortisol exposure on the ontogeny of innate immunity in rainbow trout embryos. Egg cortisol content was elevated from 4 ng/oocyte (controls - CC) to 5 (C1) and 7.5 ng/oocyte (C2) prior to fertilization (4). Lysozyme activity, intelectin, TLR-5M and TLR-5S gene expression were measured, and the intelectin and ladderlectin localization was examined in embryos using immunohistological [IH] methods. Lysozyme activity was significantly elevated in the C2 treatment group from the zygote until 13-days post fertilization (dpf), but was not affected at 21-dpf. Both intelectin and ladderlectin were present in 12-hours post-fertilization (hpf) (2-cell stage) embryos. But they had differences in response to cortisol exposure, intelectin levels (intensity of staining signal) were elevated in both cortisol treatment groups at 12-hpf and then suppressed between 36- and 48-hpf; ladderlectin was suppressed by both elevated cortisol treatments until 36-hpf compared to controls. Both lectins were distributed cell membranes and among epithelial tissues and brain tissues in late stage embryos. Intelectin mRNA was detected in oocytes and transcript levels were significantly elevated in both cortisol treatment groups relative to the controls after the 3-h incubation. There were no differences among the three treatment groups at 1- and 5-dpf, but intelectin mRNA levels were lower in the cortisol treatment groups at 13-dpf, and the transcript levels in the C2 treatment group were lower than in the CC and C1 treatment groups at 26-dpf. mRNA transcripts for the two TLR-5 genes were present in oocytes; transcript levels were significantly higher in both cortisol treatment groups relative to the controls after the 3 h incubation; TLR-5S mRNA was more abundant than that of TLR-5M mRNA. In conclusion, the ontogeny of the gene expression patterns, and the genes, lectins and lysozyme are response to increased oocyte cortisol content which indicates the importance of innate immunity in the early embryonic cells and embryo development prior to acquired immunity functions in fish. (Supported by NSERC and OMAF to JFL and JSL; NSERC Doctoral Scholarship to ML). References 1. Zapata et al. 2006. Ontogeny of the immune system of fish. Fish Shellfish Immunol. 20, 126-136; 2. Trede, et al. 2004. The use of zebrafish to understand immunity. Immunity. 20, 367-379; 3. Kindt et al. 2007. Kuby Immunology. W. H. Freeman and Co., New York, NY; 4. Li et al. 2010. The actions of cortisol in ovo on egg fertility, embryo development and growth performance of juvenile rainbow trout. Mol. Reprod. Dev. 77, 922-931; 5. Russell et al. 2008. Identification, Cloning and tissue localization of a rainbow trout (Oncorhynchus mykiss) interlectin-like protein that binds bacteria and chitin.Fish Shellfish Immunol. 25, 91-105; 6. Beutler. 2009. TLRs and innate immunity. Blood 113, 1399-1407. Keywords: cortisol, Embryo development, embryonic cell, innate immunity, ontology, rainbow trout Conference: NASCE 2011: The inaugural meeting of the North American Society for Comparative Endocrinology, Ann Arbor, United States, 13 Jul - 16 Jul, 2011. Presentation Type: Poster Topic: Developmental endocrinology Citation: Li M, Russell S, Lumsden JS and Leatherland JF (2011). In ovo elevated oocyte cortisol content induces innate immune responses in rainbow trout (Oncorhynchus mykiss) early embryonic cells. Front. Endocrinol. Conference Abstract: NASCE 2011: The inaugural meeting of the North American Society for Comparative Endocrinology. doi: 10.3389/conf.fendo.2011.04.00098 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 22 Jul 2011; Published Online: 09 Aug 2011. * Correspondence: Dr. Mao Li, University of Guelph, Biomedical Sciences, Guelph, Ontario, N1G 2W1, Canada, mao@uoguelph.ca Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Mao Li Spencer Russell John S Lumsden John F Leatherland Google Mao Li Spencer Russell John S Lumsden John F Leatherland Google Scholar Mao Li Spencer Russell John S Lumsden John F Leatherland PubMed Mao Li Spencer Russell John S Lumsden John F Leatherland Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

The ontogeny of lysozyme activity, intelectin, TLR-5M and TLR-5S gene expression and intelectin localization was examined in rainbow trout (Oncorhynchus mykiss) reared from oocytes immersed for 3h prior to fertilization in either ovarian fluid alone (CC) or cortisol-enriched ovarian fluid at either 100ngmL−1 (C1) or 1000ngmL−1 (C2) [final oocyte cortisol concentrations were ~3, ~5, and ~7.5ngoocyte−1 for the CC, C1 and C2 treatment groups, respectively]. Lysozyme activity was elevated in the cortisol-treated groups from the zygote until 13-days post fertilization (dpf), but was not affected at 21-dpf. Intelectin levels were elevated in both cortisol treatment groups at 12-hpf (2-cell stage) and then suppressed between 36- and 48-hpf. Intelectin mRNA transcript levels were elevated in both cortisol treatment groups in oocytes; there were no differences among treatment groups at 1- and 5-dpf, and suppressed in the C2 treatment group in 13-dpf and 26-dpf. TLR-5 mRNA transcripts were higher in cortisol-treated oocytes prior to fertilization; TLR-5S mRNA was more abundant than TLR-5M mRNA. The ontogeny of the gene expression patterns, and the gene, lectin and lysozyme responses to oocyte cortisol adjustments suggest an important role of innate immune systems in the early cleavage stages of embryonic cells.

Background Bisphenol A (BPA), used in the manufacture of plastics, is ubiquitously distributed in the aquatic environment. However, the effect of maternal transfer of these xenobiotics on embryonic development and growth is poorly understood in fish. We tested the hypothesis that BPA in eggs, mimicking maternal transfer, impact development, growth and stress performance in juveniles of rainbow trout (Oncorhynchus mykiss). Methodology/Principal Findings Trout oocytes were exposed to 0, 30 and 100 µg.mL−1 BPA for 3 h in ovarian fluid, followed by fertilization. The embryos were maintained in clean water and sampled temporally over 156-days post-fertilization (dpf), and juveniles were sampled at 400-dpf. The egg BPA levels declined steadily after exposure and were undetectable after 21- dpf. Oocyte exposure to BPA led to a delay in hatching and yolk absorption and a consistently lower body mass over 152-dpf. The growth impairment, especially in the high BPA group, correlated with higher growth hormone (GH) content and lower GH receptors gene expression. Also, mRNA abundances of insulin-like growth factors (IGF-1 and IGF-2) and their receptors were suppressed in the BPA treated groups. The juvenile fish grown from the BPA-enriched eggs had lower body mass and showed perturbations in plasma cortisol and glucose response to an acute stressor. Conclusion BPA accumulation in eggs, prior to fertilization, leads to hatching delays, growth suppression and altered stress response in juvenile trout. The somatotropic axis appears to be a key target for BPA impact during early embryogenesis, leading to long term growth and stress performance defects in fish.

Rainbow trout (Oncorhynchus mykiss) oocytes were incubated for 3 hr in ovarian fluid alone (CC), or cortisol-enriched ovarian fluid [100 or 1,000 ng ml(-1) (CL and CH, respectively)], after which they were fertilized; the growth and development of the embryos reared from these oocytes was monitored until first feed, and the juveniles were monitored for 9 months. The hatching rates of the CH group were significantly reduced, but the overall survival as measured at 40-week post-fertilization was similar in the three treatment groups. In addition, significant apparently biphasic changes relative to the CC group were found in the expression of some key growth-related genes in the CL and CH treatment groups, particularly IGF-1, IGF-2, GH1, GH2, GH receptors, and thyroid hormone receptors (TR alpha and TR beta). Moreover, the juveniles of the CL (but not the CH treatment group) exhibited enhanced growth; the enhanced growth could not be explained on the basis of increased feed conversion efficiency or changes in serum GH levels at the juvenile stage. Additionally, relative growth rates from the three treatment groups were similar, suggesting that the biphasic growth-enhancing effects of cortisol occurred very early in embryogenesis.

The organization, physiology and disorders of the endocrine and reproductive systems of fishes and the non-infectious factors affecting the immune system, particularly those related to the interaction of endocrine and immune system roles, are described.

This chapter discusses cortisol stress physiology and the identification of markers that will be indicative of stress and/or health effects in fish.

Quantitative RT-PCR was used to determine the profiles of expression of 10 growth- or development-related genes in rainbow trout (Oncorhynchus mykiss) embryos prior to the formation of the somatotropic (ST) axis (pituitary somatotrops and liver); embryos were sampled immediately after fertilization and water-hardening (t0), 1-h post-fertilization, and 1-, 2-, 5-, 7-, 10- and 13-days post-fertilization (dpf); expression profiles were examined in embryos reared at two temperatures (6.0 and 8.5°C), which had different developmental rates. Accumulation of mRNA encoding for GH1, GH2, IGF-1, IGF-2, two isoforms of GH receptors (GHR1, GHR2), two isoforms of IGF receptors (IGF-RIa, IGF-RIb) and two isoforms of thyroid receptor (TR), TRα and TRβ, was measured. All of these genes were expressed in the t0 samples, but the rates of expression of the different genes varied markedly. For most of the genes examined, the expression rates tended to fall within the first hour after fertilization, and remained at the lower level for between 2 and 7 days, after which there was a significant (P <0.05) and progressive increase in the number of accumulated copies of mRNA. This increase is probably associated with the commencement of embryonic genome transcription activity (EGTA), and it was generally, although not always, found later in embryos that were reared at 6.0°C compared with the faster developing embryos reared at 8.5°C. The study suggests that the EGTA begins between 2- and 5-dpf, with a staged increase in EGTA between 5- and 13-dpf.

The purpose of the study was to investigate whether dietary ration or diet composition influence the relationship between plasma growth hormone (GH) and insulin-like growth factor-1 (IGF-1) in Arctic charr (Salvelinus alpinus L.). The pattern of changes in plasma GH and IGF-1 concentrations was examined in fish fed at different ration levels (0%, 0.35% and 0.70% BW day(-1)) for 5 weeks, and in fish fed diets containing different lipid:crude protein (LCP) ratios. Ration level significantly affected plasma GH and IGF-1 concentrations; at 5 weeks the levels of both hormones in the food-deprived group were significantly lower than in fish fed the 0.70% BW day(-1) ration. Also, plasma IGF-1 levels in fish of each ration treatment group were significantly correlated with individual final body weight; no such correlation was found for GH. To examine the effects of dietary LCP ratios, fish were fed for up to 18 weeks, with one of four formulated diets that had LCP ratios (dry matter basis) of 0.35 (Diet 1), 0.43 (Diet 2), 0.51 (Diet 3) or 0.59 (Diet 4), or a commercial diet (Diet 5) which had an LCP ratio of 0.38. Statistical differences in plasma GH and IGF-1 concentrations were found only after 18 weeks. Growth hormone was significantly lower in fish fed Diets 1 and 2 compared with Diets 3 and 5, and IGF-1 was significantly lower in fish fed Diet 1 compared with Diets 2 and 5. Significant correlations between plasma GH and IGF-1 concentrations were found only for fish fed Diets 1 and 5, suggesting that the influence of diet composition on the relationship between GH and IGF-1 varies with the dietary LCP ratio in this species. The decline in plasma IGF-1 concentrations during food deprivation is similar to that described in other species; however, the unexpected decrease in plasma GH during food deprivation in this study may represent a species-specific response.

The study investigated the effects of incubation temperature, and the size of ration fed to the transitional embryo/juvenile stage of rainbow trout (Oncorhynchus mykiss) on growth, liver and gastrointestinal (GI) tract IGF-1 content, and the expression of insulin-like growth factor-related genes (IGF-1, IGF-2, IGF-RIa, and IGF-RIb) by the liver and GI tract. Embryos were reared from zygote to “swim-up” at either 8.5 °C (E8.5) or 6.0 °C (E6.0); at “swim-up” (51-days post-fertilization [dpf] and 72-dpf for the E8.5 and E6.0 groups, respectively), the embryos were transferred to grow-up tanks supplied with water at 8.5 °C. Late stage embryos (LSEs) at the same developmental stage from the two temperature treatment groups (64-dpf and 86-dpf for the E8.5 and E6.0 groups, respectively) were fed with salmonid starter diet at levels of 5.0%, 2.0%, and 0.5% of live body mass per day. Embryos were sampled just prior to first feeding (PFEs), and before complete absorption of the yolk [late stage embryos (LSEs)], and early stage juveniles (ESJs) were sampled after yolk sac absorption when they were fully reliant on exogenous sources of food. The early incubation temperature and ration levels had significant affects on mortality (with lower mortalities in the E6.0 group) and growth performance of the fish; dry body mass values for fish fed the 5.0% ration were significantly lower in the E6.0 group of LSEs and ESJs compared with the respective treatment in the E8.5 group; a similar pattern was seen for total body length, although this was only significant for the LSEs. Whole embryo IGF-1 content was significantly lower in the E6.0 group compared with the E8.5 group of PFEs, and hepatic IGF-1 content was significantly lower in the E6.0 group fed the maintenance ration (0.5%) compared with the E8.5 fed a similar ration; restricted ration significantly elevated hepatic IGF-1 content in the LSE stage for both temperature treatment groups. GI tract IGF-1 levels were considerably lower than in liver tissue, and there were no differences among treatment groups. Ration size-related differences were found for the expression of genes encoding for hepatic IGF-1, IGF-2, and IGF-RIb, and GI tract IGF-1, and IGF-2. Rearing temperature-related differences were also found for genes encoding for GI tract IGF-1, IGF-RIa, and IGF-RIb. The results of the study showed that the early rearing temperature of the embryos affected subsequent growth, and hepatic and GI tract gene expression by the LSEs and ESJs. As was the case for tissue IGF-1 content, with some exceptions, a restricted ration significantly elevated the expression of the targeted genes indicative of an important metabolic-regulating role for the IGF system during this transitional developmental phase. In addition, the higher abundance of IGF-2 mRNA compared with IGF-1 mRNA, and the higher abundance of IGF-RIa, relative to IGF-RIb, suggests that these two genes may also play a regulatory role during this transitional developmental phase.

Real time RT-PCR was used to measure the changes in the rates of synthesis of mRNA encoding for growth hormone-1 (GH1) and -2 (GH2) and insulin-like growth factor-1 (IGF-1) and -2 (IGF-2), and whole embryo GH content was measured in early stage rainbow trout (Oncorhynchus mykiss) embryos reared at two incubation temperatures (8.5 and 6.0 °C). Particular attention was paid to the phase of embryo development that preceded the appearance of the pituitary gland. GH was present in zygotes, and there were no significant changes in whole embryo GH content of the two temperature treatment groups from fertilization (t0) until the time at which GH was detectable in the pituitary gland by immunostaining. The expression of the two GH genes decreased during the first 24 h post-fertilization, and then increased significantly by 17 dpf in embryos reared at both temperatures. There was a subsequent steep increase in the number of copies of GH1 and GH2 mRNA associated with the formation of the pituitary gland evident at 23 and 34 dpf in the 8.5 and 6.0 °C groups, respectively. The number of copies of mRNA encoding for IGF-1 and IGF-2 did not change during the first 24 h post-fertilization; however, there was a significant increase in the numbers of transcripts for both genes evident by 13 dpf in embryos reared at the two incubation temperatures. The differences in the timing of the increases in GH and IGF mRNA may suggest that IGF gene expression is not GH-dependent at that stage. Moreover, the increased expression of the GH genes prior to the formation of the pituitary gland suggests that tissues other than the pituitary are expressing these genes in early embryos. The pattern of changes in GH content was similar to the pattern of GH gene expression in embryos reared at the two incubation temperatures when the age of embryos was plotted using degree-days. There were no apparent compensatory responses in GH1, GH2, IGF-1 or IGF-2 gene expression related to altered growth rates. The number of copies of IGF-2 mRNA was higher than that of IGF-1 mRNA during the early developmental period; this is consistent with the hypothesis that IGF-2 predominates during embryonic development. A differential expression of GH2 and GH1 was also observed with the overall copy numbers of GH2 mRNA being consistently higher than those of GH1.