CALCIUM INVOLVEMENT IN LUTEINIZING-HORMONE-RELEASING HORMONE-RELEASE FROM THE BOVINE INFUNDIBULUM

Bovine infundibular (stalk median eminence) explants were incubated in vitro to test the hypothesis that calcium (Ca) is involved in the release of luteinizing hormone-releasing hormone (LHRH) from LHRH neuron terminals in cattle. Right and left infundibular halves from individual heifers and/or steers were randomly assigned to either control or treated (EGTA [a Ca chelator] or verapamil [an L-type Ca channel antagonist]) groups. Each half was incubated in 600 mu l of Krebs-Ringer bicarbonate medium (KRB) in the presence or absence of a treatment agent for 180 min. At 30-min intervals, 500-mu l samples were removed from each incubate and replaced with fresh media. Spontaneous (basal) and depolarization-induced (60 mM potassium) LHRH release was evaluated by radioimmunoassay of the LHRH content in the media incubated from 91 to 120 and 121 to 150 min of culture, respectively. The effect of treatment on depolarization-induced LHRH release was analyzed by comparing the differences between spontaneous and depolarization-induced LHRH release in control and treated groups. Spontaneous LHRH release was not different between control and 1.25 mM EGTA- or 100 mu M verapamil-treated halves from steers. In contrast, steer infundibular halves incubated with EGTA (replacing Ca in KRB and chelating any Ca in the media) released less LHRH during depolarization than did control halves. In addition, verapamil-treated (to block Ca uptake by the terminal) infundibular halves from steers or heifers released less LHRH in response to depolarization than did control halves. In conclusion, these results: 1) support the hypothesis that Ca is involved in LHRH release from the bovine infundibulum, 2) suggest that the involvement of Ca may be independent of the reproductive state, and 3) demonstrate that bovine infundibular halves incubated in vitro are useful for studying selected mechanisms regulating bovine hypothalamic neurohormone release (exocytosis) from neuron terminals.