The functional location of adrenergic binding sites was studied in frog skin melanophores by injecting norepinephrine (NE) outside and inside a melanophore. In 49 groups of cells (75% of the fields tested) iontophoretic injection of NE outside the cell caused melanosome aggregation in the target cell and/or in the field. In six cells in which a resting membrane potential was measured before and after intracellular injection (10-90 nA), NE elicited no change in melanosome configuration. Once the receptors were localized, the effect of temperature on these receptors was determined by measuring the reflectance of skins (an indication of melanosome aggregation or dispersion) in two populations of frogs treated with NE, Rana pipiens pipiens (with dominant alpha-receptors) and Rana berlandieri forreri (with dominant beta-receptors). NE (0.1 mM) caused melanosome aggregation in the former and dispersion in the latter tested at 12, 22, and 40 degrees C. The iontophoretic and reflectance results suggest that the binding site of the adrenergic receptor is located on the outer surface of the plasma membrane of melanophores and that alpha- and beta-receptors evoke aggregation and dispersion, respectively, within the temperature range of these experiments.