Cholinergic Stimulation of Vasopressin Release from the Rat Hypothalamo-Neurohypophyseal System in Organ Culture

Abstract

Organ cultured hypothalamo-neurohypophyseal (HNS) explants from rats were maintained in vitro for up to 9 days. These explants contain the supraoptic nucleus with its vasopressin (AVP)-producing neurons, their axonal projections through the median eminence and axon terminals in the neurohypophysis. The presence of cells containing AVP was confirmed in 5 day cultured explants immunocytochemically. The rate of AVP release by the HNS explants remained constant over 9 days in culture. Due to simultaneous degradation of AVP by enzymes released into the culture medium by the explant, it was necessary to develop a sampling protocol which allowed simultaneous estimation of AVP release and degradation. Utilizing this protocol, the rate of AVP release by the cultured HNS was 309 .+-. 32 pg/h per explant on day 3 in culture. This rate remained essentially unchanged during the subsequent 2 days in culture. This rate of hormone release corresponds closely with other in vivo and in vitro measures of AVP secretion. Physiological responsiveness of the system was demonstrated by cholinergic stimulation of AVP release. The HNS explants were capable of a graded response to increasing doses of acetylcholine, giving a maximal 3-fold increase in AVP release over control. The cholinergic response of the HNS complex was stable on days 3-5 in culture and was blocked by hexamethonium, indicating cholinergic activation of nicotinic receptors.