Abstract
α-MSH-induced pigment dispersion in melanophores shows an absolute requirement for extracellular Ca2+. To localize Ca2+ sites involved in the mechanism of action of α-MSH we studied the effects of Ca2+ deprivation on α-MSH and forskolin-induced melanophore responses. In an in vitro melanophore system employing ventral tailfins of Xenopus tadpoles, melanophore responses were assayed in terms of pigment dispersion and the phosphorylation state of a 53 kDa melanophore-specific protein. In the same melanophore system α-MSH has been shown to specifically increase the phosphorylation of this 53 kDa protein.
Forskolin induces a dose-dependent pigment dispersion (EC50 7 × 10−7 M). In contrast to the dispersion induced by α-MSH forskolin-induced dispersion does not require extracellular Ca2+. Moreover, in a Ca2+-free medium melanophores with permanently activated MSH-receptors aggregate, but can be redispersed by the addition of forskolin. Forskolin increases 53 kDa phosphorylation in a dosedependent manner. Maximal stimulation with forskolin (10−5 M) is four-fold and equals maximal 53 kDa phosphorylation obtainable with α-MSH. The MSH-induced increase in 53 kDa phosphorylation is inhibited by Ca2+ deprivation, whereas the forskolin-induced increase is unaffected. Our results suggest that α-MSH and forskolin stimulate melanophores through a common pathway and confirm that cAMP is a second messenger in α-MSH action in this system. We conclude that the Ca2+ sites in the mechanism of α-MSH action on melanophores precede adenylate cyclase activation.