Publisher Summary This chapter describes methods for the extraction and measurement of cyclic nucleotides, their typical intracellular concentrations, and the estimation of the activities of adenylate cyclase and cAMP phosphodiesterase. Some details of the tetra- and pentaphosphates of guanosine (ppGpp and pppGpp), although not cyclic nucleotides, are also included. It is clear that studies of cyclic nucleotide levels in organisms which have not been examined previously should incorporate strict controls to demonstrate that the extracts do in fact contain the cyclic nucleotides which appear to be detected in subsequent tests, even though these tests are thought to be specific. Although cAMP and cGMP are easily identified by TLC, quantitative estimation is most easily performed by specific radioimmunoassay. The 4-fold increase in intracellular cAMP levels during nitrogen starvation of A. variabilis might suggest involvement of this cyclic nucleotide in the process of heterocyst differentiation; the constant levels observed in Synechocystis PCC 6803, a unicellular nonheterocystous strain, under the same conditions, are not inconsistent with this hypothesis. Detailed studies are required to investigate the physiological basis of the control of cAMP concentration and its potential involvement in metabolic regulation in cyanobacteria.