Rat pituitary tissue was incubated in vitro and the release of ACTH was monitored by radioimmunoassay. The release of ACTH that occurred spontaneously from unstimulated tissue was uninfluenced by a 3-hr preincubation of the tissue in the absence of calcium. ACTH release stimulated by crude hypothalamic extracts, synthetic vasopressin, dibutyryl adenosine 3’, 5’- monophosphate (cyclic AMP) and theophylline was inhibited by incubating pituitary tissue in the absence of calcium. It is concluded that in vitro activation of ACTH release, but not spontaneous ACTH release, is a calcium-dependent phenomenon. If ACTH release is mediated by cyclic AMP, then calcium ion is required for the action of cyclic AMP and need not oppose its formation.
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