Melanophore regulating effects of the melanin-concentrating hormone

During the past decade, studies on colour change mechanisms in teleost fish have revealed the existence of a second potent melanotropin, the melanin-concentrating hormone MCH. This is an hypothalamo-neurohypophysal peptide which is stored in high concentrations in the neural lobe of fish pituitaries and is released when fish adapt to a pale-coloured environment (Baker, 1988a; Kishida & Baker, 1989). Although an homologous ir. peptide is found in hypothalamic neurones of all vertebrates so far investigated, inc luding man, its distribution in animals other than the teleost is restricted mainly to the brain and only small amounts are found in the posterior pituitary gland. Moreover, with the exception of the holostean fish which are ancestral to the teleosts (Sherbrooke & Hadley, 1988), the melanophores of other vertebrates appear to lack receptors to MCH and do not respond to the peptide by melanin aggregation. Thus the use of this peptide as a colour-change hormone would seem to be an evolutionary peculiarity of certain bony fish. Indeed, in some fish such as the trout, the titre of MCH in the blood changes much more rapidly than does the a-MSH concentration (Baker, 1988b; Kishida & Baker, 1989) suggesting that MCH may have become the more important hormone for regulating physiological colour change in this species. The interest of MCH for pigment cell biologists lies in its mode of action on the melanophore, and its other involvements in pigmentation.

it is these roles which have been studied most up till now and which are reviewed briefly below. Its interest for comparative physiologists, however, is its function in the brain and how this pre-adapted it to the a role in pigmentary regulation. Our knowledge on this point is rather slight although it may well turn out to be more important than the pigmentary effects of the hormone.

Recent studies on MCH sprang from the finding that melanin-concentrating hormone, which could not be attributed to catecholamines, occurred in both the trout hypothalamus and pituitary gland, and that its abundance in both sites varied with the chromatic status of the fish (Rance and Baker, 1979; Baker & Rance, 1983). The molecule from Chum salmon pituitaries was then purified by Kawauchi et al., (1983) who showed it to be a cyclic heptadecapeptide (Fig.1). Subsequently, it was synthetized by several groups (Okamoto et al., 1984; Wilkes et al., 198 4b; Eberle et al., 1986) and it is now available commercially from Peninsula Laboratories.

The multiple effects of MCH in colour change were first apparent when it was administrated to trout over several weeks in Alzet minipumps (Baker et al., 1986). In these experiments it initially induced pallor through melanin concentration and in the long term inhibited melanogenesis This effect on melanin synthesis could be due to a direct action of MCH on the melanophores and/or to its ability to depress the release of a-MSH from the pituitary gland, which it does by a paracrine effect in the pituitary neurointermediate lobe (Barber et al., 1987). The initial rapid paling response reflects the ability of MCH to antagonize the melanin dispersing effect of a-MSH. This antagonism can be demonstrated most easily in vitro, by incubating fragments of skin or individual scales in medium containing different concentrations of MCH and a-MSH (Baker, 1988b). Such experiments show that fish species differ in their relative sensitivity to the two hormones. Thus, melanophores from the grass carp Ctenopharyngodon idellus are very responsive to low concentrations of MCH but its aggregating effect is readily overridden by equimolar concentrations of a-MSH. In contrast, although MCH is less potent on trout melanophores, it has the dominant effect when both hormones are presented together at high doses. It is possible that these species differences reflect the relative numbers of MCH and a-MSH receptors on the melanophores.

Whatever the explanation, the observations explain why it is that some fish repond to injection of fish pituitary extracts (containing both MCH and a-MSH) by melanin concentration while others respond by melanin dispersion (Pickford & Atz, 1957).

MCH appears to act on specific receptors which, in contrast to the a-MSH receptors, can be activated in the absence of extracellular calcium (Hadley et al., 1988). It is likely that a-MSH uses cAMP as its second messenger but the second messenger system of MCH is as yet un~nown. MCH is able to override both the effects of exogenous dbcAMP and the melanin dispersing effect of forskolin which increases intracellular cAMP by increasing adenyl cyclase activity ( Baker, unpub lished observations) . Nor-adrenalin from the sympathetic nervous system is also involved in causing pallor in many fish, and MCH and nor-adrenalin act synergistically on isolated trout scales (Green & Baker, in preparation ).

This suggests that they cause melanin aggregation through different second messenger systems. The active site within the MCH molecule has been studied by several groups. When fragments of salmonid MCH were tested on skin from the amazonian eel, Synbranchus, Castrucci et al., (1987) found an approximate potency of MCH1-17 (100%)= MCH5-17 > MCH1-14 (10%) > MCH5-14 (1%) These experiments reveal the importance of the C-terminal residues for receptor binding /act ivat ion, but the fact that neither MCH5-17, nor MCH5-14, appear to induce complete melanin aggregation suggests that the N-terminal sequence may also be important for full intrinsic activity in this species. Other species may show a different relative sensitivity to the fragments however, indicating species differences in receptor structure (Hadley et al., 1987; Kawazoe et al., 1987). Thus, MCH5-14 exhibits 100% potency when tested on melanophores from Tilapia mossambica (Kawazoe et al., 1987), while MCH5-17 has only about 1% potency but full intrinsic activity when tested on pigment cells of the grass carp Ctenopharvngodon (Brown et al., 1989). These studies have all used fragments of the non-homologous salmonid MCH and further studies are needed to determine whether the hormone itself has also undergone mutation in different species.

MCH causes melanin dispersion rather than aggregation when tested at high concentrations on melanophores of amphibians and reptiles (Wilkes et al., 1984a, Baker et al., 1985; Ide et al., 1985), and it also has this effect in vitro on skin of the amazonian eel (Castrucci et al., 1987) low doses of the peptide causing melanin aggregation and high doses melanin dispersion. The dispersive effect is not observed if calcium is absent from the extracellular medium, nor is it exhibited by the fragments MCH5-17 and MCHS5-14 (Hadley et al., 1988) which, as noted above, exhibit melanin concentrating activity. Hadley and co-workers have therefore concluded that melanin dispersion results from an interaction between the N-terminal region of MCH with the a-MSH receptors, even, though there is no apparent similarity between the primary structures of a -MSH and MCH (Fig. 1).

Such an interpretation could explain why the melanin aggregating potency of MCH is enhanced in the absence of extracellular calcium (Hadley et al., 1988a; Baker, unpublished). MCH has a similar MSH-like activity on mouse B-16 melanoma cells, inducing tyrosinase activity when added to the culture medium at a concentration of 10-6M (Baker et al., 1985a).

The non-pigmentary effects of MCH are still poorly understood. Besides depressing the release of a -MSH from the pars intermedia, MCH will also inhibit the release of ACTH from the corticotrophs so that fish kept in white tanks become less responsive to moderate stresses (Gilham & Baker, 1985). MCH delivered by Alzet minipump has the same effect on ACTH release (Baker et al., 1986) and this is partly due to a direct action on the pituitary pars distalis (Baker et al., 1985b) although effects at the hypothalamic level are also possible. Other effects of MCH in the brain remain to be explored but recent studies in rats show that it is able to partially antagonize certain behavioural effects induced by iv injections of a -MSH (dee Graan et al., 1989). It seems likely that the pigmentary role of MCH evolved either from its inhibitory interaction with a -MSH in the brain or from its ability to depress the release of a-MSH from the pituitary gland.

References
Baker BI (1988a) : In "The Melanotropins", Vol. II, pp. 159-174 (ed. ME. Hadley) CRC Press Inc.
Baker BI (198 8b) In "Advances in Pigment Cell Research", Proc. 13th International Pigment Cell Conference, pp 505-516 (ed. JT. Bagnara) Alan R. Liss Inc.
Baker BI et al (1985) : J Endocr 106:R5-R8.
Baker BI et al (1986) : Gen Comp Endocr 63:62-69.
Baker BI et al (1985) : Peptides 6:1125-1130.
Baker BI, Rance TA (1983) : Gen Comp Endocr 50:423-431.
Barber LD et al (1987) : Gen Comp Endocr 65:79-86.
Brown DW et al (1989) : Biopolymers, submitted.
Castrucci AM de L et al (1987) : Life Sci 40:1845-1851.
Eberle AN et al (1986) : J Chem Soc Perkin Trans 1:361-367.
Gilham ID, Baker BI (1985) : J Endocr 105:99-105.
Graan PNE et al (1989) : Peptides, in press.
Hadley ME et al (1988) In "Advances in Pigment Cell Research", Proc. 13th International Pigment Cell Conference, pp 531-545 (ed. JT Bagnara) Alan R. Liss Inc.
Hadley ME et al (1987) : Life Sci 40:1139-1145.
Ide H et al (1985) : Gen Comp Endocr 58:486-490.
Kawauchi H et al (1983) : Nature (London) 305:321-333.
Kawazoe I et al (1987) : Int J Pept Prot Res 29:714-721.
Kishida M et al (1989) : Gen Comp Endocr 73, in press.
Okamoto et al (1984) : Chem Parm Bull 32:2963-2970.
Pickford GE, Atz J (1957) : The physiology of the pituitary gland of fishes. Zoological Soc, NY.
Rance TA, Baker BI (1979) : Gen Comp Endocr 37:64-73.
Sherbrooke WC, Kadley ME (1988) : Pigment Cell Res 1:344-349.
Wilkes BC et al (1984a) : Science 224:1111-1113.
Wilkes BC et al (1984b) : Biochem Biophys Res Comm 122:613-619.

Fig. 1

The primary structures of MCH and a -WSH

MCH H-Asp-Thr-Met -Arg-Cys-Met-Val -Gly-Arg -Va l-Tyr-Arg-Pro-Cys -Trp-Glu-Val-OH

MSH Ser-Tyr-Ser-Met -Glu-His-Phe-Arg -Trp-Gly-Lys-Pro-Va 1-NH2

 

Bridget I. Baker
School of Biological Sciences
Bath University
Bath - Avon BA2 7AY, UX
Phone : 0225-826826 ext. 5871