To clarify the mechanism of OT secretion of hypothalamus and pituitary origin, in vitro, the guinea pig hypothalamo-neurohypophyseal complex (HNC) was utilized in this study. The HNC, including paraventricular and supraoptic nuclei, was removed and maintained in Eagle's Minimal Essential Medium (pH 7.4) for 7 days at 37 degrees C in a 95% air 5% CO2 environment. The OT content in the hypothalamus and the pituitary remained constant during the period of culture of the HNC. Synthetic OT, hypothalamus or pituitary extracts were eluted separately by ion exchange chromatography and the elution patterns obtained were similar. The stimulation of the HNC with 100mM KCl in the culture caused an increase in OT release. These results suggest that the cultured HNC can be a useful in vitro model for studies of the OT secretion mechanism. After adding several substances to the culture medium, the amount of OT release from the HNC into the culture medium over a 20 minute period, was measured by our own radioimmunoassay. The addition of prostaglandin F2 alpha to the HNC resulted in an increase in OT release while stimulation of D2 or E2 failed to increase. Stimulation of the HNC with vasoactive intestinal polypeptide (VIP) produced an increase in OT release, suggesting the existence of a positive feedback effect of VIP on OT release.