Regulation of corticotropin-releasing factor-binding protein expression in amygdalar neuronal cultures.
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 microM), 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 microM 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.
Kasckow
J W
JW
Cincinnati VAMC, Psychiatry Service, Cincinnati, OH 45220, USA. Jkasckow@pol.net
Regmi
A
A
Seasholtz
A F
AF
Mulchahey
J J
JJ
D020121:5' Untranslated Regions / Q000235:genetics; D000679:Amygdala / Q000166:cytology*; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics*; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology; D005260:Female; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D005938:Glucocorticoids / Q000494:pharmacology; D015850:Interleukin-6 / Q000502:physiology; D009474:Neurons / Q000737:chemistry / Q000166:cytology / Q000502:physiology*; D011247:Pregnancy; D011493:Protein Kinase C / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013312:Stress, Physiological / Q000503:physiopathology
D020121:5' Untranslated Regions; D002352:Carrier Proteins; D005938:Glucocorticoids; D015850:Interleukin-6; D012333:RNA, Messenger; C067133:corticotropin releasing factor-binding protein; D003907:Dexamethasone; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C