Regulation of corticotropin-releasing factor-binding protein expression inamygdalar neuronal cultures

Corticotropin-releasing factor-binding protein (CRF-BP) is known to regulat e 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 syst em (CNS) limbic and cortical areas, including the amygdala. The signal tran sduction pathways which regulate amygdalar CRF-BP are not well understood. In this report, we have examined the effect of protein kinase A and C activ ators, CRF, dexamethasone and interleukin-6 (IL6) on CRF-BP mRNA and protei n expression in dissociated fetal amygdalar cultures. CRF-BP mRNA levels we re determined by Northern analysis following 12 h treatment with the follow ing agents: forskolin (1-30 mu M), CRF (1-1000 nM), phorbol-12-myristate-13 -acetate (TPA; 1-50 nM), dexamethasone (1-100 nM) and IL6 (10-500 pM). Sign ificant 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 Weste rn analysis of total cellular protein after treatment with the same agents. Twenty-four hour treatment with 30 mu M forskolin, 1000 nM CRF, 50 nM TPA, 100 pM IL6 or 100 nM dexamethasone significantly increased CRF-BP expressi on (P < 0.05, n=3 for each treatment). The primary cultures were then trans fected with a rat CRF-BP-reporter construct containing 3500 base pairs of C RF-BP 5' flanking DNA. Treatment with all five agents produced statisticall y 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 whi ch may play a significant role in promoting behavioural changes.