Effect of forskolin and exogenously administered oxytocin mRNA on oxytocinrelease by dispersed hypothalamic cultures

Differential vasopressin (VP) gene expression and oxytocin (OT) gene expres sion were observed in hypothalamic cultures derived from 14-day-old rat fet uses, with VP but not OT being induced by treatment with forskolin and 3-is obutyl-1-methylxanthine. These cultures were used to demonstrate that exoge nous VP mRNA could be taken up and translated into releasable VP. In the cu rrent studies a similar culture preparation was used to test the hypothesis that, due to the similarity in the mRNA and prohormone structures of VP an d OT, the VP-expressing neurons in the cultures would be capable of utilizi ng exogenous OT mRNA for synthesis of releasable OT. Although OT release wa s increased by the administration of exogenous OT mRNA, endogenous OT gene expression was also observed. To determine what had induced OT gene express ion in the current cultures, the undefined components of the culture prepar ation, e.g., the glial feeder layer and the serum component of the culture medium, were evaluated. Restraining growth of the glial carpet with cytosin e-arabinoside did not alter OT gene expression. Use of a defined medium sup plemented with B-27 induced optimal OT gene expression. From this, it is po ssible 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 s erum-free, B-27-supplemented medium may provide a useful model system for s tudying OT gene regulation. (C) 2001 Academic Press.