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1 Biology, University of Waterloo, Waterloo, Ontario, Canada
* To whom correspondence should be addressed. E-mail: mvijayan{at}sciborg.uwaterloo.ca.
We used primary cultures of trout hepatocytes and a physiological dose of cortisol (100 ng ml-1), mimicking stressed levels in salmonid fish, to address the impact of glucocorticoid stimulation on glucocorticoid receptor (GR) mRNA abundance and protein content. Cortisol significantly elevated GR mRNA content over a 24 h period; this increase was abolished by actinomycin D suggesting transcriptional control of GR. However, cortisol significantly decreased GR protein content leading us to hypothesize that lower GR protein content may be regulating GR mRNA abundance. Indeed, treatment of hepatocytes with MG-132, a proteasomal inhibitor shown to prevent GR degradation by cortisol, abolished cortisol-mediated GR mRNA upregulation. Also, geldanamycin, an hsp90- specific inhibitor, abolished the GR mRNA increase evident with cortisol, but did not modify cortisol-induced increases in abundance of mRNA for PEPCK, a glucocorticoid-responsive gene, or hepatocyte glucose release. Together, our results suggest a negative feedback loop for GR gene regulation by cortisol in trout hepatocytes. The autoregulation of GR may be a crucial step in the physiological stress response process, especially in modulating energy dependent processes that are glucocorticoid-dependent, including gluconeogenesis.
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