Regulation of Hypothalamic Corticotropin-Releasing Hormone Transcription by Elevated Glucocorticoids

Abstract

Negative glucocorticoid feedback is essential for preventing the deleterious effects of excessive hypothalamic pituitary adrenal axis axis activation, with an important target being CRH transcription in the hypothalamic paraventricular nucleus. The aim of these studies was to determine whether glucocorticoids repress CRH transcription directly in CRH neurons, by examining glucocorticoid effects on glucocorticoid receptor (GR)-CRH promoter interaction and the activation of proteins required for CRH transcription. Immunoprecipitation of hypothalamic chromatin from intact or adrenalectomized rats subjected to either stress or corticosterone injections showed minor association of the proximal CRH promoter with the GR compared with that with phospho-CREB (pCREB). In contrast, the Period-1 (Per1, a glucocorticoid-responsive gene) promoter markedly recruited GR. Stress increased pCREB recruitment by the CRH but not the Per1 promoter, irrespective of circulating glucocorticoids. In vitro, corticosterone pretreatment (30 minutes or 18 hours) only slightly inhibited basal and forskolin-stimulated CRH heteronuclear RNA in primary hypothalamic neuronal cultures and CRH promoter activity in hypothalamic 4B cells. In 4B cells, 30 minutes or 18 hours of corticosterone exposure had no effect on forskolin-induced nuclear accumulation of the recognized CRH transcriptional regulators, pCREB and transducer of regulated CREB activity 2. The data show that inhibition of CRH transcription by physiological glucocorticoids in vitro is minor and that direct interaction of GR with DNA in the proximal CRH promoter may not be a major mechanism of CRH gene repression. Although GR interaction with distal promoter elements may have a role, the data suggest that transcriptional repression of CRH by glucocorticoids involves protein-protein interactions and/or modulation of afferent inputs to the hypothalamic paraventricular nucleus.

Methylation at the CpG island shore region upregulatesNr3c1promoter activity after early-life stress

“…This unanticipated finding concurs with another recent report. 45 Together with the present observation that GR-mediated negative feedback is less efficient (delayed) in ELS mice (Fig. S8), these findings suggest that GR regulation of the Crh gene (specifically, in contrast to Fkbp, Sgk1, and Dusp1) is persistently altered by ELS, the molecular underpinnings of which warrant future analysis.…”

“…The lack of effects on Crh are consistent with the results of other studies showing that basal or stimulated Crh transcription in vivo are barely affected by alterations of the corticosterone milieu over a broad range of concentrations. 45 The parvocellular division of the PVN harbors distinct sub-populations of peptidergic neurons, among them neurons that express only Crh or Avp, or both. [46][47][48] Here we used triple fluorescence immunohistochemistry to identify which subsets of PVN neurons express GR.…”

Molecular Regulation of Corticotropin-Releasing Hormone Gene Expression in Parvocellular Neurons of the Hypothalamic Paraventricular Nucleus

“…Third, glucocorticoids do not directly affect cyclic AMP induced CREB phosphorylation and nuclear accumulation of the co-activator TORC in vitro [52]. Fourth, GR immunoprecipitation in hypothalamic chromatin from rats injected with corticosterone show a lack of GR recruitment by the CRH promoter, in spite of marked recruitment by the glucocorticoid-dependent, period 1 gene [52]. Previous reports have shown that a conserved sequence upstream of the essential À247 CRE is capable of binding GR in gel shift assays and mediates glucocorticoid dependent repression in reporter gene assays [54,55].…”

“…Second, injection of high doses of corticosterone at the time of stress exposure does not prevent the increases in CRH hnRNA induced by stress [53]. Third, glucocorticoids do not directly affect cyclic AMP induced CREB phosphorylation and nuclear accumulation of the co-activator TORC in vitro [52]. Fourth, GR immunoprecipitation in hypothalamic chromatin from rats injected with corticosterone show a lack of GR recruitment by the CRH promoter, in spite of marked recruitment by the glucocorticoid-dependent, period 1 gene [52].…”

“…Glucocorticoids also inhibit CRH transcription but most evidence indicates that in physiological conditions indirect mechanisms mediate this effect [51,52]. First, suppression of the glucocorticoid surge by adrenalectomy does not affect the declining phase of stress induced CRH hnRNA, indicating that factors other than glucocorticoids are responsible for the rapid decline of CRH transcription during persistent stress.…”