Various substances were tested for an effect on MSH release from the pituitary of the lizard, Anolis carolinensis. On the basis of the following findings it is believed that serotonin may be an MSH releasing factor in this animal. 1. (1) Serotonin creatinine sulphate in the incubation medium stimulated MSH release from hemipituitaries at all stages of the incubation. At 2.6 × 10 −8 M serotonin, a stimulatory effect could just be detected, while at 7.8 × 10 −8 M, total MSH released was increased twofold and at 2.6 × 10 −7 M, more than threefold. 5-Methoxytryptamine at 2.6 × 10 −7 M doubled MSH release, but at the same concentration 5-hydroxytryptophan had no stimulatory effect. The effect of 2.6 × 10 −7 M serotonin was reduced by imidazole (10 m M), suggesting that serotonin acts to stimulate MSH release through the adenyl cyclase-cyclic AMP mechanism. 2. (2) Subcutaneous injections of serotonin creatinine sulphate darkened green Anolis. Injected doses ranging from 3 to 10 μg/g caused skin darkening, beginning after 2 min and lasting for about 15 min. Since it lacked a direct darkening effect in vitro on melanocytes of Anolis skin, injected serotonin was believed to be acting directly on the pars intermedia of the pituitary. 3. (3) The precursor of serotonin, 5-hydroxytryptophan, had no direct action on melanocytes nor on the release of MSH from the pituitary in vitro, but injections of 14–200 μg/g caused darkening of lizards, beginning after a short delay of about 12 min and lasting for up to 48 hr with the larger doses. Tryptophan injections did not cause lizards to darken. Melatonin injections (20–100 μg/g) reversed 5-hydroxytryptophan-induced darkening, an effect which lasted for about 30 min with the larger doses of melatonin. Melatonin, by itself, in vitro, had no clear effect on MSH release from the pituitary but the stimulatory effect of 2.6 × 10 −7 M serotonin was reduced significantly by the presence of equimolar melatonin. Injected synthetic mammalian MRF (H.cysH. tyr. ile. gln. asn. OH) did not cause darkening of lizards and MIF-I (H-pro, leu. gly. NH 2) did not cause paling of 5-hydroxytryptophan-darkened lizards. MIF-I and MRF at 1 μg/ml had no effect on MSH release in vitro.