The effect of various substances on parathyroid hormone (PTH) release was studied in vitro, in an attempt to determine whether adenosine 3′,5′–monophosphate (cyclic AMP) may be involved in regulating PTH release. Bovine parathyroid gland slices were incubated for 5 hr in tissue culture medium. The medium was changed each hour and analyzed for PTH concentration by radioimmunoassay. After the first 2 hr of incubation in medium with 1.5 mM calcium (Ca) concentration, the composition was changed to that of the test condition. Incubation with low Ca (0.75 mM) medium increased PTH release, and incubation with high Ca (3.0 mM) medium decreased PTH release significantly. PTH release was increased significantly by 10 mM dibutyryl cyclic AMP, 2 mM aminophylline, or 0.45 mM epinephrine. These in vitro observations a) strongly suggest that PTH release is influenced by cyclic AMP in the parathyroid cell, and b) further suggest that the known hypocalcemic stimulus of PTH release is mediated by the adenyl cyclasecyclic AMP system. (Endocrinology92: 687, 1973)