Integumental reddish-violet coloration owing to novel dichromatic chromatophores in the teleost fish, Pseudochromis diadema.

In the reddish-violet parts of the skin of the diadema pseudochromis Pseudochromis diadema, we found novel dichromatic chromatophores with a reddish pigment and reflecting platelets. We named these novel cells 'erythro-iridophores'. In standard physiological solution, erythro-iridophores displayed two hues, red and dark violet when viewed with an optical microscope under ordinary transmission light and epi-illumination optics, respectively. Under transmission electron microscopy, however, we observed no typical red chromatosomes, i.e., erythrosomes, in the cytoplasm. High-performance thin-layer chromatography (HPTLC) analysis of the pigment eluted from the erythro-iridophores indicated that carotenoid is the main pigment generating the reddish color. Furthermore, when the irrigating medium was a K(+)-rich saline solution, the color reflected from the erythro-iridophores changed from dark violet to sky blue, but the red coloration remained. The motile activities of the erythro-iridophores may participate in the changes in the reddish-violet shades of the pseudochromis fish.

Dermal matrix proteins initiate re-epithelialization but are not sufficient for coordinated epidermal outgrowth in a new fish skin culture model.

We have established a new culture system to study re-epithelialization during fish epidermal wound healing. In this culture system, fetal bovine serum (FBS) stimulates the epidermal outgrowth of multi-cellular layers from scale skin mounted on a coverslip, even when cell proliferation is blocked. The rate of outgrowth is about 0.4 mm/h, and at 3 h after incubation, the area occupied by the epidermal sheet is nine times larger than the area of the original scale skin. Cells at the bottom of the outgrowth show a migratory phenotype with lamellipodia, and "purse string"-like actin bundles have been found over the leading-edge cells with polarized lamellipodia. In the superficial cells, re-development of adherens junctions and microridges has been detected, together with the appearance and translocation of phosphorylated p38 MAPK into nuclear areas. Thus, this culture system provides an excellent model to study the mechanisms of epidermal outgrowth accompanied by migration and re-differentiation. We have also examined the role of extracellular matrix proteins in the outgrowth. Type I collagen or fibronectin stimulates moderate outgrowth in the absence of FBS, but development of microridges and the distribution of phosphorylated p38 MAPK are attenuated in the superficial cells. In addition, the leading-edge cells do not have apparent "purse string"-like actin bundles. The outgrowth stimulated by FBS is inhibited by laminin. These results suggest that dermal substrates such as type I collagen and fibronectin are able to initiate epidermal outgrowth but require other factors to enhance such outgrowth, together with coordinated alterations in cellular phenotype.

Bhc-cNMPs as either water-soluble or membrane-permeant photoreleasable cyclic nucleotides for both one- and two-photon excitation.

Cyclic nucleoside monophosphates (cNMPs) play key roles in many cellular regulatory processes, such as growth, differentiation, motility, and gene expression. Caged derivatives that can be activated by irradiation could be powerful tools for studying such diverse functions of intracellular second messengers, since the spatiotemporal dynamics of these molecules can be controlled by irradiation with appropriately focused light. Here we report the synthesis, photochemistry, and biological testing of 6-bromo-7-hydroxycoumarin-4-ylmethyl esters of cNMP (Bhc-cNMP) and their acetyl derivatives (Bhc-cNMP/Ac) as new caged second messengers. Irradiation of Bhc-cNMPs quantitatively produced the parent cNMPs with one-photon uncaging efficiencies (Phiepsilon) of up to one order of magnitude better than those of 2-nitrophenethyl (NPE) cNMPs. In addition, two-photon induced photochemical release of cNMP from Bhc-cNMPs (7 and 8) can be observed with the two-photon uncaging action cross-sections (delta(u)) of up to 2.28 GM (1 GM=10(-50) cm(4) s photon(-1)), which is the largest value among those of the reported Bhc-caged compounds. The wavelength dependence of the delta(u) values of 7 revealed that the peak wavelength was twice that of the one-photon absorption maximum. Bhc-cNMPs showed practically useful water solubility (nearly 500 microM), whereas 7-acetylated derivatives (Bhc-cNMPs/Ac) were expected to have a certain membrane permeability. Their advantages were demonstrated in two types of biological systems: the opening of cAMP-mediated transduction channels in newt olfactory receptor cells and cAMP-mediated motility responses in epidermal melanophores in scales from medaka fish. Both examples showed that Bhc and Bhc/Ac caged compounds have great potential for use in many cell biological applications.