The Effects of Ions, Metabolic Inhibitors, and Colchicine on Luteinizing Hormone-Releasing Hormone Release from Superfused Rat Hypothalami*

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Abstract
In the present study, the effects of ions, metabolic inhibitors, and colchicine on the release of immunoreactive LHRH from superfused male rat hypothalami were examined. High extracellular potassium reliably stimulated LHRH release; this K+-evoked release has been attributed to membrane depolarization triggering influx of Na+ and Ca++ and leading to exocytosis of LHRH. As such, stimulated release of LHRH was clearly Ca++ dependent. However, in the absence of Na+ or in the presence of the Na+ channel blocker tetrodotoxin, K+-stimulated release persisted, indicating that Na+ was not obligatory in this release process. Ouabain, a Na+-K+-APase inhibitor, blocked K+-stimulated release of LHRH, although basal release was unaffected. 2,4-Dinitrophenol, which uncouples oxidative phosphorylation, surprisingly resulted in significant elevations in both basal and K+-stimulated LHRH release, whereas metabolic energy inhibitors, cyanide and iodoacetic acid, reduced basal release and completely blocked the K+-stimulated release of LHRH. K+-evoked release of LHRH is thought to involve a microtubular transport mechanism, since colchicine (10-6 or 10-5 M) significantly inhibited this release in a dose-dependent manner. However, colchicine at a concentration of 10-4 M dramatically increased basal and K+-stimulated LHRH release as well as hypothalamic LHRH content by an unknown mechanism. These results strongly suggest that intracellular Ca++ plays a key role in the active release process of this neuropeptide from hypothalamic fragments perifused in vitro. It can also be postulated that the microtubular transport mechanism is an important component of this neurosecretory process.