CALCIUM AND CAMP-MEDIATED STIMULATION OF FOS IN CULTURED HYPOTHALAMICTYROSINE HYDROXYLASE-IMMUNOREACTIVE NEURONS

Immediate-early genes, such as c-fos, couple extracellular signals to genetic changes in the cell. We have previously demonstrated that depo larization with 50 mM KCl increases Fos immunoreactivity in hypothalam ic tyrosine hydroxylase (TH) and oxytocin immunoreactive (-ir) neurons in primary culture. This Fos activation occurs within 1.5-2 h in TH-i r cells. In the present study, we examined the effects of depolarizati on, glutamate receptor activation and adenylyl cyclase stimulation on Fos-ir to determine the possible mechanism(s) of Fos activation in TH- ir neurons. Hypothalamic cultures were treated with KCl, glutamate or forskolin, and Fos and TH were visualized immunocytochemically. Forsko lin increased the percentage of Fos/TH-ir neurons in a dose-dependent manner, with a maximal stimulation of 53.4 +/- 4.5% Fos/TH-ir neurons at 30 mu M forskolin. The dose-response curve for glutamate was steep, with a maximal stimulation of 24.8 +/- 2.1% Fos-ir neurons at 100 mu M. 50 mM KCl resulted in 50.0 +/- 0.8% Fos/TH-ir neurons. Pretreatment with verapamil decreased KCl induced Fos-ir by 57%, glutamate by 65% and forskolin by 39%. Combined drug administration demonstrated signif icant additivity between forskolin and glutamate, and forskolin and KC l, however, no significant additivity was found with KCl and glutamate . The results are discussed in terms of cAMP and calcium mediation of the Fos response to these stimuli.