While ACTH is known to induce skin pigmentation in man, its effects on cultured human melanocytes have not been investigated. Using a culture system free of artificial mitogens, we report for the first time that ACTH stimulates melanogenesis in cultured human melanocytes. While ACTH, alpha-MSH and the synthetic alpha-MSH analogue Nle4DPhe7 alpha-MSH all stimulate the activity of tyrosinase, the rate limiting enzyme in melanogenesis, and all produce a 50% increase in the melanin content of the cells at a concentration of 10(-8)-10(-7) mol/l, the shapes of the dose response curves differ: those for the MSH peptides are sigmoidal while those for ACTH are biphasic. In addition, human melanocytes are able to respond to concentrations of ACTH comparable with physiological plasma levels. We suggest that ACTH may be relatively more important than alpha-MSH as a pigmentary hormone in man and could have a physiological role in skin pigmentation.