Seasonal effects of GnIH on basal and GnRH-induced goldfish somatotrope functions
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Figure 1
The effect of gGnIH on serum GH (upper panel) and pituitary gh mRNA levels (lower panel) during early recrudescence (October) in vivo. Goldfish received two injections of 2 μg gGnIH, at time 0 and 12 h; fish that had received injections of PBS served as controls. Pituitaries and blood samples for serum collection were removed at 2 h following the second injection. Serum GH values were analyzed by RIA (mean±s.e.m.; n=12). Abundance of pituitary transcripts was determined by qPCR. gh mRNA values were normalized against β-actin mRNA and expressed relative to those for controls (mean±s.e.m.; controls, n=6); control groups were normalized to 100%. Different letters denote statistically significant differences (ANOVA followed by Tukey's test, P<0.05). Tissues that did not resemble the appropriate gonadal stage were not used.
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Figure 2
The effect of gGnIH on serum GH (upper panel) and pituitary gh mRNA levels during mid-recrudescence (December) in vivo. Goldfish received two injections of 2 μg gGnIH, at time 0 and 12 h; fish that had received injections of PBS served as controls. Pituitaries and blood samples for serum collection were removed at 2 h following the second injection. Serum GH values were analyzed by RIA (mean±s.e.m.; n=12). Abundance of pituitary transcripts was determined by qPCR. gh mRNA values were normalized against β-actin mRNA and expressed relative to those for controls (mean±s.e.m.; controls, n=6); control groups were normalized to 100%. Different letters denote statistically significant differences (ANOVA followed by Tukey's test, P<0.05). Tissues that did not resemble the appropriate gonadal stage were not used.
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Figure 3
The effect of gGnIH on serum GH (upper panel) and pituitary gh mRNA (lower panel) levels during late recrudescence (March) in vivo. Goldfish received two injections of 2 μg gGnIH at time 0 and 12 h; fish that had received injections of PBS served as controls. Pituitaries and blood samples for serum collection were removed at 2 h following the second injection. Serum GH values were analyzed by RIA (mean±s.e.m.; n=14). Abundance of pituitary transcripts was determined by qPCR. gh mRNA values were normalized against β-actin mRNA and expressed relative to those for controls (mean±s.e.m.; controls, n=7); control groups were normalized to 100%. Different letters denote statistically significant differences (ANOVA followed by Tukey's test, P<0.05). Tissues that did not resemble the appropriate gonadal stage were not used.
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Figure 4
The dose-related effects of gGnIH on GH release (upper panel) and gh mRNA (lower panel) levels in primary cultures of dispersed goldfish pituitary cells prepared from fish at early gonadal recrudescence (October). After 12 h of static treatment, a medium sample was removed for GH level analysis via RIA, and the RNA from cells was extracted for determination of transcript abundance. Gh transcript abundance was determined by qPCR and the results normalized against β-actin and expressed as a percentage of control values (mean±s.e.m.; n=8 from two independent cell preparations). Different letters denote statistically significant differences (ANOVA followed by Tukey's test, P<0.05).
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Figure 5
The dose-related effects of gGnIH on GH release (upper panel) and gh mRNA (lower panel) levels in primary cultures of dispersed goldfish pituitary cells prepared from fish at mid-gonadal recrudescence (December). After 12 h of static treatment, a medium sample was removed for GH level analysis via RIA, and the RNA from cells was extracted for determination of transcript abundance. gh transcript abundance was determined by qPCR and the results normalized against β-actin and expressed as a percentage of control values (mean±s.e.m.; n=8 from two independent cell preparations). Different letters denote statistically significant differences (ANOVA followed by Tukey's test, P<0.05).
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Figure 6
The dose-related effects of gGnIH on GH release (upper panel) and gh mRNA (lower panel) levels in primary cultures of dispersed goldfish pituitary cells prepared from fish at late gonadal recrudescence (March). After 12 h of static treatment, a medium sample was removed for GH level analysis via RIA, and the RNA from cells was extracted for determination of transcript abundance. gh transcript abundance was determined by qPCR and the results normalized against β-actin and expressed as a percentage of control values (mean±s.e.m.; n=8 from two independent cell preparations). Different letters denote statistically significant differences (ANOVA followed by Tukey's test, P<0.05).
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Figure 7
Effects of gGnIH (10 nM) on the GH response to (A) sGnRH (100 nM) and (B) cGnRHII (100 nM) in perifused dispersed goldfish pituitary cells prepared from goldfish of mixed sex at mid-recrudescence (December–February). gGnIH was applied as a 5-min pulse (arrow), 25 min into a 1-h treatment with GnRH (horizontal bar). GH content in perifusates collected in 5-min fractions was analyzed by RIA. GH values for each treatment column were normalized as a percentage of pretreatment values (average of the first five fractions collected before hormone treatment). Net GH responses to GnRH were quantified as ‘area under the curve with baseline subtracted’ (duration of quantification indicated by the vertical dotted lines; baseline defined as the average of the three fractions before the quantification period). GH release profiles are presented in the left panels and quantified net GH responses to GnRH are presented in the right panels (mean±s.e.m., n=8 from four separate cell preparations). Different letters denote significant differences between treatment groups (ANOVA followed by Tukey's test, P<0.05).
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Figure 8
Effects of gGnIH (10 nM) on the GH response to (A) sGnRH (100 nM) and (B) cGnRHII (100 nM) in perifused dispersed goldfish pituitary cells prepared from goldfish of mixed sex at late recrudescence/sexually matured stages (May–early June). GnRH was applied as a 5-min pulse (arrow), 25 min into a 1-h treatment with gGnIH (horizontal bar). GH content in perifusates collected in 5-min fractions was analyzed by RIA. GH values for each treatment column were normalized as a percentage of pretreatment values (average of the first five fractions collected before hormone treatment). Net GH responses to GnRH were quantified as ‘area under the curve with baseline subtracted’ (duration of quantification indicated by the vertical dotted lines; baseline defined as the average of the three fractions before the quantification period). GH release profiles are presented in the left panels and quantified net GH responses to GnRH are presented in the right panels (mean±s.e.m., n=8 from four separate cell preparations). Different letters denote significant differences between treatment groups (ANOVA followed by Tukey's test, P<0.05).
- © 2014 Society for Endocrinology
