Evidence for synergism between copper and prostaglandin E 2 in stimulating the release of gonadotropin-releasing hormone from median eminence explants: Na +/Cl − requirements

Abstract

Copper (Cu) and PGE 2 are known to stimulate LHRH release from expiants of the median eminence area (MEA) by two mechanisms distinguishable by their Ca 2+ dependence. Moreover, exposure to Cu and PGE 2 results in an amplified release of LHRH which is partially Ca 2+ dependent, thus, resembling the release process stimulated by PGE 2 alone. We have shown that LHRH release stimulated by Cu alone is Na +/Cl − dependent. By defining the Na +/Cl − dependence of PGE 2- and Cu/PGE 2-stimulated release of LHRH, we wished to ascertain if there is synergism between Cu and PGE 2 actions. MEA of adult male rats were incubated for 5 min with 150 μM Cu and then for 15 min with 10 μM PGE 2 (Cu/PGE 21). Controls were incubated with Cu or PGE 2. LHRH release into the medium was evaluated by RIA. Substituting Cl − in the incubation buffer with the non-permeant anion, isethionate, did not alter PGE 2 stimulation of LHRH release, but it drastically inhibited Cu/PGE 2 stimulation of LHRH release, indicating that this process requires a permeant monovalent anion. PGE 2 and Cu/PGE 2 stimulation of LHRH release were both inhibited when Na + was substituted with Li +, or when 0.5 mM ouabain was included in the Na +-containing buffer; neither 10 μM tetrodotoxin (TTX) nor 100 μM amiloride were inhibitory. To ascertain if Na + is required for Cu uptake, we evaluated the uptake of 67Cu by MEA expiants and found that neither ouabain nor Li + inhibited uptake, indicating that the extracellular Na + and the activity of Na +/K + ATPase are required for the process of LHRH release. In summary, (i) Cu and PGE 2 stimulation of LHRH release are distinct processes, based on their Ca 2+ and Cl − requirements; and (ii) the process of Cu/PGE 2 stimulation of LHRH release exhibits the ionic requirements of both secretagogues. This is consistent with Cu and PGE 2 synergistically acting on the LHRH neuron, which results in amplification of the release processes initiated by each of the secretagogues.