Potent and cooperative feedback inhibition of adenylate cyclase activity by calcium in pituitary-derived GH 3 cells

Abstract

Calcium (Ca 2+) ion concentrations that are achieved intracellularly upon membrane depolarization or activation of phospholipase C stimulate adenylate cyclase via calmodulin (CaM) in brain tissue. In the present study, this range of Ca 2+ concentrations produced unanticipated inhibitory effects on the plasma membrane adenylate cyclase activity of GH 3 cells. Ca 2+ concentrations ranging from 0.1 to 0.8 μM exerted an increasing inhibition on enzyme activity, which reached a plateau (35–45% inhibition) at around 1 μM. This inhibitory effect was highly cooperative for Ca 2+ ions, but was neither enhanced nor dependent upon the addition of CaM (1 μM) to EGTA-washed membranes. The inhibition was greatly enhanced upon stimulation of the enzyme by vasoactive intestinal peptide (VIP) and/or GTP. Prior exposure of cultured cells to pertussis toxin did not affect the inhibition of plasma membrane adenylate cyclase activity by Ca 2+, although in these membranes, hormonal (somatostatin) inhibition was significantly attenuated. Maximally effective concentrations of Ca 2+ and somatostatin produced additive inhibitory effects on adenylate cyclase. The addition of phosphodiesterase inhibitors demonstrated that inhibitory effects of Ca 2+ were not mediated by Ca 2+-dependent stimulation of a phosphodiesterase activity. These observations provide a mechanism for the feedback inhibition by elevated intracellular Ca 2+ levels on cAMP-facilitated Ca 2+ entry into GH 3 cells, as well as inhibitory crosstalk between Ca 2+-mobilizing signals and adenylate cyclase activity.