Effect of forskolin and exogenously administered oxytocin mRNA on oxytocin release by dispersed hypothalamic cultures.
Differential vasopressin (VP) gene expression and oxytocin (OT) gene expression were observed in hypothalamic cultures derived from 14-day-old rat fetuses, with VP but not OT being induced by treatment with forskolin and 3-isobutyl-1-methylxanthine. These cultures were used to demonstrate that exogenous VP mRNA could be taken up and translated into releasable VP. In the current studies a similar culture preparation was used to test the hypothesis that, due to the similarity in the mRNA and prohormone structures of VP and OT, the VP-expressing neurons in the cultures would be capable of utilizing exogenous OT mRNA for synthesis of releasable OT. Although OT release was increased by the administration of exogenous OT mRNA, endogenous OT gene expression was also observed. To determine what had induced OT gene expression in the current cultures, the undefined components of the culture preparation, e.g., the glial feeder layer and the serum component of the culture medium, were evaluated. Restraining growth of the glial carpet with cytosine-arabinoside did not alter OT gene expression. Use of a defined medium supplemented with B-27 induced optimal OT gene expression. From this, it is possible to conclude that the components included in B-27 are sufficient for OT gene expression.Factors included in earlier lots of sera may have been responsible for suppression of OT gene expression. Cultures maintained in serum-free, B-27-supplemented medium may provide a useful model system for studying OT gene regulation.
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Department of Physiology and Biophysics, Finch University of Health Sciences/The Chicago Medical School, 3333 Green Bay Road, North Chicago, Illinois 60064, USA.
Sladek
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D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology*; D005333:Fetus; D005786:Gene Expression Regulation / Q000187:drug effects*; D007031:Hypothalamus / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D010121:Oxytocin / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors; D014158:Transcription, Genetic
D012333:RNA, Messenger; D005576:Colforsin; D010121:Oxytocin