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Elevations of inducible cAMP early repressor (ICER), the repressor isoform of the cAMP-responsive element modulator (CREM), are associated with protein binding to the corticotrophin releasing hormone (CRH) promoter and termination of CRH transcriptional responses to stress. To determine whether endogenous ICER production represses CRH transcription, we examined the effect of CREM siRNA on forskolin-stimulated ICER formation and CRH transcription in the hypothalamic cell line, 4B, and in primary cultures of hypothalamic neurons. Cotransfection of 4B cells with CREM siRNA and a CRH promoter-driven luciferase reporter gene markedly reduced the induction of ICER by forskolin and potentiated the stimulatory effect of forskolin on CRH promoter activity, compared with cells cotransfected with a nonspecific oligonucleotide. The role of ICER on endogenous CRH expression was studied in primary cultures of hypothalamic neurons by examining the effect of CREM siRNA on forskolin-induced primary transcript (CRH hnRNA) using intronic real-time PCR. As observed during stress in vivo, forskolin-stimulated CRH hnRNA was transient, increasing up to 60 min and declining to near basal values by 3 h. Transfection of CREM siRNA reduced forskolin-induced ICER by about 45% 48-h later and partially reversed the declining phase of CRH hnRNA production at 3 h. The data provide evidence that endogenous ICER formation is required for termination of CRH transcription and support the hypothesis that ICER is part of an intracellular feedback mechanism limiting the activation of CRH transcription during stress.

Cyclic AMP Inducible Early Repressor Mediates the Termination of Corticotropin Releasing Hormone Transcription in Hypothalamic Neurons