Potentiation of proopiomelanocortin gene expression in cultured pituitary cells by benzodiazepines.
BACKGROUND
Benzodiazepines are frequently used not only as a part of general anesthesia but also for the purpose of sedation during regional anesthesia. Effects of these drugs on the hypothalamic-pituitary-adrenal axis activity have been studied, but are still controversial. It is not known whether benzodiazepines affect expression of proopiomelanocortin, precursor protein of adrenocorticotropic hormone and related peptides.


METHODS
AtT20PL cell line, a clone of AtT20/D16v mouse corticotroph tumor cells stably transfected with approximately 0.7 kilobases (kb) of the rat proopiomelanocortin 5' promoter-luciferase fusion gene, was used. In the presence or absence of diazepam or midazolam, cells were stimulated by corticotropin-releasing hormone (CRH) or forskolin. Proopiomelanocortin gene expression was estimated by measurement of luciferase activity. Furthermore, to study the mechanism of benzodiazepine effects, cyclic adenosine 3',5'-monophosphate (cyclic AMP) efflux was measured by enzyme immunoassay.


RESULTS
Diazepam and midazolam dose-dependently increased the proopiomelanocortin gene expression induced by CRH or forskolin. The potentiating effect was not affected by benzodiazepine receptor antagonists flumazenil and PK11195, but was abolished by a cyclic AMP-dependent protein kinase inhibitor H89. Cyclic AMP efflux induced by CRH or forskolin was also enhanced by diazepam and midazolam. In the presence of isobutylmethylxanthine, a nonspecific phosphodiesterase inhibitor, potentiation of proopiomelanocortin gene expression and enhancement of cyclic AMP efflux by benzodiazepines were not observed.


CONCLUSIONS
Benzodiazepines potentiate the effect of CRH or forskolin on proopiomelanocortin gene expression. The potentiating effect is not mediated by the benzodiazepine receptors, but its mechanism probably involves inhibition of phosphodiesterase.
Fukuda
Kazuhiko
K
Department of Anesthesia, Kyoto University Hospital, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan. kfukuda@kuhp.kyoto-u.ac.jp
Uetsuki
Nobuo
N
Uga
Hisatoshi
H
Hashiguchi
Mitsuko
M
Sato
Masami
M
Hisano
Taizo
T
Segawa
Hajime
H
Iwasaki
Yasumasa
Y
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D014151:Anti-Anxiety Agents / Q000494:pharmacology*; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D003975:Diazepam / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D007700:Kinetics; D008156:Luciferases / Q000235:genetics; D051379:Mice; D008874:Midazolam / Q000494:pharmacology; D011333:Pro-Opiomelanocortin / Q000235:genetics*; D014162:Transfection
D014151:Anti-Anxiety Agents; D005576:Colforsin; D011333:Pro-Opiomelanocortin; D008156:Luciferases; D003975:Diazepam; D008874:Midazolam; D015056:1-Methyl-3-isobutylxanthine