INCREASED VASOPRESSIN SECRETION FROM HYPOTHALAMIC CULTURES FOLLOWING ADMINISTRATION OF EXOGENOUS VASOPRESSIN MESSENGER-RNA

In Brattleboro rats, exogenous vasopressin (VP) mRNA can be accumulate d, transported, and translated by magnocellular neurons. To determine whether this phenomenon may also occur in magnocellular neurons of nor mal rats, dispersed hypothalamic neuronal cultures of fetal Sprague-Da wley rats were exposed to VP mRNA. The cultures were maintained in eit her control medium or medium containing the cAMP elevating drugs, IBMX (3-isobutyl-1-methylxanthine), and forskolin for 23 or 30 days of cul ture to induce VP synthesis and secretion. Following removal of the IB MX and forskolin at Day 23, VP secretion into the medium declined to b aseline by 30 days in vitro, but administration of VP mRNA to these cu ltures on Day 30 resulted in a 5-fold increase in VP content of the me dium (P = 0.005) after 6 h and a 2.5-fold increase after 24 h (P = 0.0 02). Administration of VP mRNA to the cultures treated continuously wi th IBMX and forskolin also resulted in a small increase in VP secretio n which did not reach significance after either 6 or 24 h. When cultur es prepared with continuous I/F were exposed to antisense VP mRNA, VP secretion into the media was decreased by 58%, and VP immunoreactive p erikayra were difficult to observe. This demonstrates that the increas e in VP release observed after the addition of sense VPmRNA did not re flect a nonspecific effect of the addition of mRNA to the culture medi um. Autoradiography of cultures administered H-3- or P-32-VP mRNA for 24 h revealed silver grains associated with varicosities, perikarya, a nd neuritic processes of neurophysin (NP)-positive, but not NP-negativ e neurons. These results suggest that exogenously administered mRNA ha s access to cell translation systems in cultured hypothalamic neurons as well as magnocellular neurons of Brattleboro rats. (C) 1996 Academi c Press, Inc.