Chronic dibutyryl cAMP (dbcAMP) treatment was observed not only to potentiate the differentiating actions of nerve growth factor (NGF) in PC12 cells, but to render them completely dependent on trophic support for survival even in the presence of serum proteins. When both NGF and dbcAMP were withdrawn from doubly differentiated PC12 cultures, degenerative events occurred after a lag period of 12-18 h, and by 48 h < 5-10% of the cells remained viable. Reduction in [3H]dopamine uptake, an index of cell function and neurite integrity, paralleled cell demise. At the cellular level, approximately 20-30% of the nuclei exhibited clear signs of chromatin fragmentation, as characterized by propidium iodide staining, suggesting that degeneration occurred by apoptosis. The cells could be rescued completely from degeneration by dbcAMP or by other cAMP analogues, whereas NGF and depolarization were also effective, but only partially. Phorbol 12-myristate-13-acetate failed to afford protection. If deprivation was interrupted, cell demise could be stopped by restoration of initial culture conditions. Degenerative changes produced by deprivation and recovery processes were not inhibited by macromolecular synthesis inhibitors, e.g. cycloheximide and actinomycin-D. However, chronic addition of cycloheximide prior to deprivation greatly impaired the differentiation of NGF/dbcAMP cells, allowing these cells to withstand trophic support withdrawal. Altogether our results indicate that the cAMP transduction pathway plays a crucial role not only in the differentiation but also in the survival of NGF/dbcAMP PC12 cells.