In this review the conformational studies of natural enkephalins (H-Tyr-Gly-Gly-Phe-Met-OH; the [Met(5)]enkephalin and H-Tyr-Gly-Gly-Phe-Leu-OH; the [Leu(5)]enkephalin), their acyclic and cyclic analogues, including those carried out in our laboratory, performed by experimental and theoretical methods and their combination, are described. Emphasis is given on the role of conformational constraints introduced by cyclization on activity at the micro and delta opioid receptors. Comparison of the conformations of cyclic enkephalin analogues with high delta-receptor activity with those of potent rigid non-peptide delta-receptor agonists indicates that the proximity of the aromatic side chains in positions 1 and 4 as well as the N-terminal amino group is desirable for the activity at the delta opioid receptors; early conformational studies also suggested that spatial separation of the aromatic side chains and rigidity of the cyclic backbone is desirable for micro-receptor activity. The results of our recent conformational studies performed with the use of fluorescence and NMR spectroscopy as well as theoretical calculations indicate, however, that these structural features are not necessary for activity at the micro opioid receptors. Methods applied to the determination of the conformation of flexible peptides, such as Nuclear Magnetic Resonance (NMR), fluorescence spectroscopy, and theoretical conformational analysis are also discussed briefly.