Abstract

C-6 astrocytoma cells which were incubated with norepinephrine for 3 hr lost the ability to respond to the hormone by accumulating high concentrations of adenosine cyclic 3',5'-monophosphate (cAMP). The altered response of the treated cells was not attributable to a greater release of cAMP from the cells into the incubation fluid. Comparison of the rates of cAMP accumulation by and disappearance from intact cells provided evidence that cells treated with norepinephrine hydrolyzed cAMP more rapidly than did naive cells. Increased cyclic nucleotide phosphodiesterase activity was found in homogenates of treated cells. The increase in phosphodiesterase activity was prevented by cycloheximide and actinomycin D, indicating that an induction of new enzyme synthesis was responsible for the increase in enzyme activity. The increased phosphodiesterase activity was apparent with cAMP but not with guanosine cyclic 3',5'-monophosphate (cGMP) as substrate in the enzyme assay, suggesting that the induced form of the enzyme did not hydrolyze cGMP. The increased enzyme activity did not require Ca²⁺ for activity. Cycloheximide only partially prevented the effect of norepinephrine treatment on cAMP accumulation by intact cells. Further experimentation revealed that the norepinephrine-stimulated adenylate cyclase was decreased in cells treated with norepinephrine in the presence of cycloheximide.