Stereo high voltage electron microscopy of melanophores : Matrix transformations during pigment movements and the effects of cold and colchicine

Pigment migration in isolated melanophores of the angelfish, Pterophyllum scalare, has been studied by high voltage electron microscopy. Cells were isolated from the scales by collagenase and allowed to spread on Formvar and carbon-coated gold grids. Melanophores were then fixed by glutaraldehyde and osmium tetroxide and critical-point dried for viewing of whole cells in a high voltage electron microscope (1000 kV). The three-dimensional organization of the cytoplasmic matrix was stereoscopically examined in different states of pigment distribution, as well as under cold and colchicine treatment. The most prominent matrix constituent is an extensive mesh of cytoplasmic filaments (microtrabeculae) 2–18 nm in diameter that make contact to microtubules, pigment granules, and mitochondria. Microtrabeculae undergo dramatic changes in structural appearance in association with different phases of pigment movements. Cells fixed in the process of pigment aggregation are characterized by thickened and beaded trabeculae which may form irregular clots. Part of this material trails behind centripetally moving melanosomes. In dispersing cells, microtrabeculae are straight and of relatively uniform thickness throughout their length and form a highly ordered three-dimensional lattice. Reconstruction of the mesh in part precedes the arrival of pigment granules.Under the influence of cold or colchicine treatment, microtrabeculae show a high degree of polymorphism, being beaded, branched, or flattened with globose ends. Rather formless heaps are found associated with the surface of pigment granules. Since, however, these treatments also remove microtubules, the other important component of the cytoplasmic frame, alterations in microtrabecular structure may simply be mediated through removal of this organelle. In an attempt to separate the effects on microtrabeculae and microtubules from one another, cells have been cold-treated for only 15 min, a procedure that leaves a considerable portion of microtubules intact. Also under these conditions, microtrabeculae are beaded or transformed to globose heaps and flattened sheets.The observations suggest an involvement of microtrabeculae in the process of granule movement. Centripetal melanosome migration thereby seems associated with a collapse of microtrabeculae which again are reconstructed during pigment dispersion. The cold and colchicine experiments indicate direct effects of these agents on the structure and possibly also the function of the trabecular mesh. The significance and possible chemical composition of microtrabeculae is discussed.