The addition of physiological concentrations of either cAMP or cGMP stimulated the release of RNA from isolated prelabeled rat liver nuclei to a fortified cytosol in a cell-free system. The released RNA was shown to be primarily mRNA by its binding to oligo(dT)-cellulose and its sedimentation profile. Treatment of rats with cAMP or cGMP 30 min prior to the preparation of cyclic nucleotides on the cell-free system. Cyclic nucleotides stimulation of RNA release occurred in systems prepared from resting rat liver, Novikoff hepatoma, and Morris hepatoma 5123D, but not the 18-h regenerating liver. The response of the cell-free system to added cyclic nucleotides reflected the in vivo concentration of these substances in the tissues from which the system was prepared. Those with high in vivo levels were not stimulated while those with lower levels did respond to added cyclic nucleotides. Neither cAMP nor cGMP had an appreciable effect on rRNA release.