Biomedical Applications of Synthetic Melanotropins

Alpha-melanotropin (alpha-melanocyte stimulating hormone, alpha-MSH) is a hormone produced by the pituitary gland of most vertebrate animals. This melanotropic peptide, Ac-Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Val-NH2, regulates melanin pigmentation of the skin of some mammals. Although MSH may be absent from the human pituitary gland, this peptide can stimulate pigment formation in human skin. We have synthesized several analogues of alpha-MSH, which are superpotent, prolonged-acting, and resistant to inactivation by serum enzymes. One such analogue, [NLe⁴, D-Phe⁷]alpha-MSH, has proven particularly useful in a number of physiological studies. In addition, some [Nle⁴, D-Phe⁷]-substituted fragment analogues of MSH are even more active than the native hormone, alpha-MSH. For example, these analogues are 100–1,000 times more active than alpha-MSH in stimulating S-91 mouse melanoma tyrosinase activity in vitro. We have successfully labeled one such peptide to high specific activity; this melanotropin, [³H]-Ac-[Nle⁴, D-Phe⁷]alpha-MSH_4–11NH₂, has been shown by others to bind to B16 melanoma cells. We have also conjugated several ligands (fluorescein and biotin) to [Nle⁴, D-Phe⁷]alpha-MSH. These melanotropin conjugates might prove useful for melanotropin receptor studies and for the clinical localization of metastatic melanoma. We have demonstrated that [Nle⁴, D-Phe⁴]alpha-MSH can be topically applied and transdermally delivered across the skin of mice and humans in vitro, as determined by bioassay and RIA. Initial toxicologic studies indicate that the analogue is nontoxic to mice and is not mutagenic. Studies are underway to determine whether this analogue may prove useful as a “tanning hormone” for increasing the pigmentation of light-skinned individuals or possibly even for treating people with certain hypopigmentary disorders.     

The Control of Color in Birds

SYNOPSIS. The colors of birds result from deposition of pigments—mainlymelanins and carotenoids—in integumentary structures,chiefly the feathers. The plumages of birds indicate their age,sex, and mode of living, and play important roles in camouflage,mating, and establishment of territories. Since feathers aredead structures, change of color of feathers is effected throughdivestment (molt) and replacement. The color and pattern ofa feather are determined by the interplay of genetic and hormonalinfluences prevailing in its base during regeneration. Mostbirds replace their feathers at least once annually. Some wearthe same kind of basic plumage all the time butothers alternatea basic and breeding plumage, either in one (the male) or bothsexes. Still others may have more than two molts, adding supplementalplumage at certain times in the plumage cycle. The varietiesof patterns of molt, the kinds of plumage, and the colors andpatterns of feathers among birds apparently are the result ofseveral kinds of selection pressures working through evolution.     

Physiological Color Changes in Reptiles

SYNOPSIS. The physiological regulation of color changes in reptilesas studied in the lizard, Anolis carolinensis, is discussed.In Anolis, the ability to adapt to a background is dependentupon the level of circulating MSH, therelease of which is dependenton information received through the eyes. Blinded (or intact)lizards are brown under conditions of strong illumination andgreen under conditions of lower light intensities, and, again,these color changes are regulated by MSH. According to Kleinholz,color changes in the blinded lizard are regulated by dermalphotoreceptors. High or low temperatures directly affect thecolor of Anolis skins and alter the rate at which skins respondto hormones. Aggregationof melanin granules within Anolis melanophoresin response to sympathomimetic stimulation is regulated throughalpha adrenergic receptors whereas dispersion of melanin granulesin response to such stimulation is controlled through beta adrenergicreceptorspossessed by the melanophores. Most Anolis melanophores possessboth alpha and beta adrenergic receptors, but some melanophorespossess only beta adrenergic receptors. In the normal physiologyof the lizard, under conditions of stress, stimulation of alphaadrenergic receptors by catecholamines leads to an "excitement—pallor"followedby an "excitement—darkening" resulting from stimulationof beta adrenergic receptors which causes dispersion of melaningranules within localized populations of melanophores. Thus,in Anolis, dispersion of melanin granules within melanophoresis regulated by both MSH and by catecholamines. Evidence ispresented that the intracellular level of cyclic 3', 5'-AMPwithin melanophores may be responsible for the regulation ofmovement of melanin granules.     

Evidence for a Calcium/Calmodulin Involvement in Density-Dependent Melanogenesis in Murine B16 Melanoma Cells

Previous work from our laboratory has shown that both cyclic AMP and calcium/calmodulin appear to be involved in the regulation of melanogenesis in murine B16 melanoma cells. In these cells as in murine Cloudman S91 cells, melanogenic responsiveness to melanocyte-stimulating hormone (MSH) varies with cell density in culture. Our objective in this study was to learn more about the intracellular systems involved in the control of melanogenesis, particularly the role played by calcium. The melanogenic response to αMSH was compared to the response to drugs affecting intracellular free calcium and calmodulin over a range of cell densities in B16F1 cells. αMSH-stimulated melanin production was extremely density-dependent but αMSH-stimulated cyclic AMP production was independent of cell density. The melanogenic response to agents that increased intracellular calcium (A23187) or inhibited intracellular calmodulin varied with cell density. A drug (TMB8) that lowered intracellular free calcium, however, increased melanogenesis independently of cell density. At high cell density it was found that an elevation in calcium decreased melanogenesis, whereas agents that reduced calcium or inhibited calmodulin activity increased melanogenesis. At low cell density, however, the inhibitory response to A23187 was lost and in some experiments even stimulated melanogenesis. These data suggest that the calcium/calmodulin signalling system has an inhibitory influence on melanogenesis, and its expression, which depends upon cell density, may also modulate the response to stimulatory agents such as αMSH.     

Development of Amblyospora campbelli (Microsporida: Amblyosporidae) in the Mosquito Culiseta incidens (Thomson)

The complete life cycle of Amblyospora campbelli (Kellen and Wills, 1962) (Microsporida: Amblyosporidae) requires a two-host system involving the mosquito host, Culiseta incidens (Thomson), and an obligatory intermediate copepod host. The parasite has dimorphic spore development producing meiospores (haploid condition) and binucleated spores (diploid condition), either as an exclusive infection or simultaneously (within females only). This is the 1st known report of concurrent spore development within an adult mosquito host, and, therefore, shows the Amblyospora campbelli system to be uniquely different from other Amblyospora spp. cycles previously described. The significance of dimorphic spore development is discussed. In females, diplokaryotic meronts may invade oenocytes, causing a benign-type of infection. A blood-meal is required to initiate sporulation of the binucleate spore. The binucleate spore contains the sporoplasm involved in transovarial transmission. A 2nd sporulation sequence, primarily in adipose tissue, may involve both males and females. In this sequence, repeated merogonic division greatly increased the density of diplokaryotic meronts and generally involved most of the body of the host. Production of meiospores, unlike that for the binucleate spore, appeared to be spontaneous (i.e. no obligatory blood meal). Survivorship of male and female larval mosquitoes was nearly equal. Adult females spread the parasite in three ways: transovarial, transovum, and by meiospore deposition.     

Coccidia of Brazilian Mammals: Eimeria corticulata N. Sp. (Apicomplexa: Eimeriidae) from the Anteater Tamandua tetradactyla (Xenarthra: Myrmecophagidae) and Eimeria zygodontomyis N. Sp. from the Cane Mouse Zygodontomys lasiurus (Rodentia: Cricetidae)

Feces from a specimen of Tamandua tetradactyla (Linn.) from Portel, Para State, north Brazil, contained two different coccidial oocysts; one identified as Eimeria tamanduae Lainson 1968, and the other as a new species, described here as Eimeria corticulata n. sp. Oocysts of E. corticulata are ellipsoidal, 37.4 × 30.4 (31.2–43.7 × 23.7–35.0) μm, shape index (length/width) 1.2 (1.0–1.5). Oocyst wall 2.5–3.7 μm thick and composed of two layers; an outer thick, brown-yellow one with radial striations, and a thin inner smooth one: no visible micropyle. Oocyst residuum a large globule of about 10.7 × 10.3 μm, usually accompanied by a number of smaller attached globules. Sporocysts ellipsoidal, 21.0 × 11.0 (20.0–22.5 × 10.0–12.5) μm, with a conspicuous Stieda body: shape index 1.9 (1.6–2.2). Sporocyst residuum a small number of scattered granules: sporozoites 18.7 × 5.0 μm, with a large posterior refractile body. Eimeria zygodontomyis n. sp. is described in feces from Zygodontomys lasiurus (Lund) from the Serra dos Carajas, Para. Oocysts ellipsoidal to cylindrical, 16.5 × 12.0 (13.7–18.7 × 11.2–12.3) μm, shape index 1.4 (1.2–1.5). Wall colorless, smooth, single-layered and about 0.6 μm thick: no micropyle. No oocyst residuum, but a polar granule of about 1.8 × 1.0 μm is sometimes present. Sporocysts ellipsoidal, 8.4 × 5.5(7.5–8.7 × 5.0–6.2) μm, shape index 1.5 (1.4–1.7), with a thin colorless wall and a delicate Stieda body. Sporozoites enclose a compact residuum of about 2.5 × 3.7 μm.