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Regulation of Corticotropin‐Releasing Factor‐Binding Protein Expression in Amygdalar Neuronal Cultures
Author(s) -
John Kasckow,
Ajit Regmi,
Audrey F. Seasholtz,
J.J. Mulchahey
Publication year - 1999
Publication title -
journal of neuroendocrinology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.062
H-Index - 116
eISSN - 1365-2826
pISSN - 0953-8194
DOI - 10.1046/j.1365-2826.1999.00413.x
Subject(s) - medicine , forskolin , endocrinology , corticotropin releasing hormone , dexamethasone , urocortin , protein kinase c , messenger rna , amygdala , phorbol , glucocorticoid , protein kinase a , biology , signal transduction , hypothalamus , chemistry , receptor , kinase , gene , microbiology and biotechnology , biochemistry
Corticotropin‐releasing factor‐binding protein (CRF‐BP) is known to regulate the bioavailability of CRF and may also play a role in stress behaviours. CRF‐BP has been localized in the pituitary as well as central nervous system (CNS) limbic and cortical areas, including the amygdala. The signal transduction pathways which regulate amygdalar CRF‐BP are not well understood. In this report, we have examined the effect of protein kinase A and C activators, CRF, dexamethasone and interleukin‐6 (IL6) on CRF‐BP mRNA and protein expression in dissociated fetal amygdalar cultures. CRF‐BP mRNA levels were determined by Northern analysis following 12 h treatment with the following agents: forskolin (1–30 μM), CRF (1–1000 nM), phorbol‐12‐myristate‐13‐acetate (TPA; 1–50 nM), dexamethasone (1–100 nM) and IL6 (10–500 pM). Significant increases in CRF‐BP mRNA were observed in response to forskolin (30 mM), CRF (100, 1000 nM), IL6 (100, 500 pM), TPA (50 nM) and dexamethasone (100 nM; P<0.05 for all; n=3–6 for all). We extended our observations of CRF‐BP expression to the protein level by performing semiquantitative Western analysis of total cellular protein after treatment with the same agents. Twenty‐four hour treatment with 30 μM forskolin, 1000 nM CRF, 50 nM TPA, 100 pM IL6 or 100 nM dexamethasone significantly increased CRF‐BP expression (P<0.05, n=3 for each treatment). The primary cultures were then transfected with a rat CRF‐BP‐reporter construct containing 3500 base pairs of CRF‐BP 5′ flanking DNA. Treatment with all five agents produced statistically significant increases above control (P<0.05; n=3 for each). The results suggest that CRF‐BP in the amygdala is stimulated by numerous pathways which may play a significant role in promoting behavioural changes.