Rolipram is a clinically effective antidepressant with selective cAMP phosphodiesterase (PDE) inhibiting properties. (+/-)-[3H]Rolipram binds with high affinity (Kd = 2.52 +/- 0.47 nM) to sections of rat brain (Hill number = 0.90 +/- 0.05). Binding is stereospecific. Association of (+/-) [3H]rolipram to sections is rapid (47% of specific binding in the first minute, kobs = 0.52 min-1). Dissociation of (+/-)-[3H]rolipram exhibits non first order kinetics (3 component model; t1/2 = 2.5 min, 50 min and 6 h, respectively). A number of PDE inhibitors reduce (+/-)-[3H]rolipram binding to the level of nonspecific binding ((-)-rolipram, IC50 = 0.9 nM; (+/-)-rolipram, IC50 = 1.5 nM; Ro 20-1724, IC50 = 11 nM; ICI 63.197, IC50 = 35 nM; medazepam, IC50 = 240 nM; diazepam, IC50 = 1200 nM; IBMX, IC50 = 3800 nM). In vitro autoradiography reveals high binding site densities in the cerebellum, olfactory bulb, lateral septal nucleus, frontal cortex, subiculum and CA1 of hippocampus. Most of the labeled structures are part of the limbic system. In vivo autoradiography of (+/-)-[3H]rolipram binding shows much more nonspecific binding than in vitro, nevertheless the distribution pattern of (+/-)-[3H]rolipram binding sites is similar. A comparison of the distribution pattern of (+/-)-[3H]rolipram binding sites with that of an antidepressant (monoamine oxidase inhibitor, monoamine uptake inhibitor) reveals no overlap. Limited, though significant correlations exist with the distribution of beta 1-adrenergic, adenosine1 and glutamate/quisqualate receptors as well as protein kinase C, but not with beta 2-adrenergic receptors and forskolin binding sites.