The teleost cone cytoskeleton. Localization of actin, microtubules, and intermediate filaments.

This laboratory has been using the teleost retinal cone as a model for studying the mechanisms and regulation of retinal cell motility. In previous inhibitor studies, the authors have shown that dark-induced cone elongation requires microtubules, whereas light-induced contraction requires actin filaments. This study examines the distributions of actin filaments, microtubules, and intermediate filaments in the cone cytoskeleton. Actin filaments have been localized in isolated cones by labeling with fluorescent derivatives of phalloidin; microtubules were localized by immunofluorescent labeling with anti-tubulin. Actin, microtubule, and intermediate filament distributions have also been examined in detergent-lysed motile cell models of cones fixed with a new method that enhances preservation of the cytoskeleton. Longitudinal bundles of actin filaments extend from the cone's calycal processes through the ellipsoid and into the myoid. No actin filaments are detectable in the perinuclear region and axon, but filaments are present in both pre- and post-synaptic components of the synapse. Intermediate filaments are numerous in the perinuclear region and cone axon but relatively sparse in the myoid. In contrast, microtubule distribution is more uniform: numerous longitudinally oriented microtubules are present throughout the length of the cell. Thus the cone cytoskeleton reflects the highly polarized shape and function of the cell, with actin filaments localized to the distal movable part of the cell and intermediate filaments localized to the proximal part of the cell, which is anchored in the retina.

Lymphoid organs of teleost fish. III. Splenic lymphoid tissue of Rutilus rutilus and Gobio gobio.

The ultrastructure of splenic lymphoid tissue of two teleost fish, Rutilus rutilus and Gobio gobio, constitutes discrete foci around the small arteries and "melano-macrophage" centres. It contains small and medium lymphocytes, lymphoblasts, macrophages and plasma cells. Plasmacytopoiesis and macrophage-lymphocyte clusters have been described and the significance of "melano-macrophage" centres and macrophage-lymphocyte clusters is discussed.

[Electron microscopic characteristics of neurosecretory cells in the preoptic nucleus of sturgeon fry]. / Elektronnomikroskopicheskaia kharakteristika neirosekretornykh kletok preopticheskogo iadra mal'kov osetra

In the sturgeon fry nucleus preopticus (Npo), light and weakly differentiated neurosecretory cells (NSC) as well as differentiated (i-cells) were revealed. Among these types of cells, there were the transitional forms. No "picnomorphic" cells were found in any Npo zone. NSC contained different amounts of elementary secretory granules 110-270 nm in diameter. The distribution of organelles was evaluated statistically, their number increasing from i-cells to the NSC. These data suggest that growth and differentiation of the NSC take place during this stage of ontogeny. The results are discussed in terms of specific functioning and maturity of the NSC.

Determinants of directional specificity in the regeneration of lamprey spinal axons.

The projection patterns of regenerating spinal axons in the larval sea lamprey (Petromyzon marinus) were determined by intracellular injection of HRP. Four hundred and eighty-six of 562 stained axons and axon-like neurites (87%) arising from Muller and Mauthner axons, giant interneurons, and dorsal cells terminated in an orientation similar to that of their counterpart control cells. Therefore, lamprey spinal axons regenerate selectively along their normal projection paths. During the first 4 weeks of recovery, i.e., before any had regenerated beyond the transection site, 91 of 114 axons and long neurites (80%) projected in the proper direction. Thus, the correctness of the final projection patterns did not result from selective retraction of randomly directed long neurites. When the cords were doubly transected 1 cm apart, orientation of regenerating neurites remained normal both within the 1 cm island and in the adjacent spinal cord. This suggests that the directional specificity of axonal regeneration was determined neither by the location of the scar nor by the availability of channels formed by the degenerating fibers. Finally, removing 1 cm of spinal cord eliminated potential synaptic targets for regenerating axons on either side of the lesion, but did not affect the direction of axonal growth. These findings are consistent with the hypothesis that the regeneration of lamprey spinal axons is guided by local chemical cues that persist long after the pathways are formed early in development.

Histochemical and biochemical studies of carbonic anhydrase activity in the opercular epithelium of the euryhaline teleost, Fundulus heteroclitus.

Carbonic anhydrase (CAH) activity was biochemically measured and histochemically localized (at both the light and electron microscope levels) in isolated opercular membranes from teleost fish, Fundulus heteroclitus, adapted to freshwater (FW), seawater (SW), and double-strength seawater (2 x SW). The normal morphology of this membrane showed that its epithelial portion consisted of five cell types: (1) chloride cells, which have been previously implicated as responsible for the active chloride transport across the epithelium; (2) mucous cells; (3) pavement cells, which formed the major portion of the free epithelial surface; (4) supportive cells, which had an abundance of intermediate (10 nm)-type filaments suggesting a structural role for these cells; and (5) vesicular cells, which were characterized by various types of membrane-bound vesicles, including lysosomes, and numerous free ribosomes. Vesicular cells may be stem cells and/or endocrine cells. Hansson's histochemical method for CAH revealed cobalt sulfide reaction product confined to the following structures in fish from each environment: (1) chloride cells: throughout the cytoplasm and some nuclear staining; (2) mucous cells: throughout the cytoplasm, some nuclear staining, and some in mucous granules; (3) vesicular cells: confined to lysosomes, some of the vesicles, and nucleoli; (4) a small portion of the intracellular space between adjacent vesicular cells and supportive cells; and (5) supportive cells: in nucleoli and occasionally in larger membrane-bound lysosomelike structures. Acetazolamide (10(-5) M) and potassium cyanate (KCNO) (10(-1) M) in Hansson's incubation medium completely inhibited the formation of reaction product. Biochemical determination of CAH activity on vascularly perfused, isolated opercular membranes showed no statistically significant difference in enzyme activity between environmental groups. The following units of activity/mg opercular membrane protein were measured: FW: 0.63 +/- 0.02; SW: 0.43 +/- 0.08; 2 x SW: 0.64 +/- 0.09.

Cell rearrangement and directional migration in pronephric duct development.

The morphology of the directed migration of the pronephric duct rudiment of three vertebrates, the salamander, chick and sturgeon, has been examined by scanning electron microscopy. Of particular interest in this paper are the morphology of the duct tip, the role of cell rearrangement, and the relation of duct extension to somite segmentation. The duct rudiments of all three species have motile cell processes (lamellipodia and filopodia) largely confined to their posterior tips. The salamander and sturgeon embryos extend their duct rudiments by extensive cell rearrangements. A short, wide rudiment is elongated to form a long, thin one. The chick duct rudiment stays about the same width and apparently gains volume by cell proliferation. The salamander duct rudiment's posterior tip is always two somites behind the last formed somite. Both the sturgeon and chick embryo's duct rudiments lie well posterior of the last segmented somite adjacent to segmental plate mesoderm. There is still a close coupling, however, between the posterior progression of the duct rudiments and the advancing wave of somite segmentation.

Paths of axons in the visual system of perciform fish and implications of these paths for rules governing axonal growth.

The optic nerve of many perciform fish is ribbon-shaped, and axons from ganglion cells in specific parts of the retina are consistently found in specific places in this ribbon. I utilized this organization to fill selected groups of axons with horseradish peroxidase. I then traced these groups of axons through the nerve and across the tectum to their terminal arbors. The paths of the axons suggest that axons use a number of different mechanisms to guide them to their correct terminal sites. At some points they appear simply to grow along the surface created by earlier axons, but at other points they seem to be using cues more complex than simple mechanical guidance. In addition, I have demonstrated that for every anulus of ganglion cells on the retina there is an anulus of terminal arbors on the tectum. With time the terminals in a given anulus must move caudally to keep the retinotopic map centered on the tectum while the tectum continues growing nonsymmetrically . I have shown both that the anuli of terminals do remain roughly centered on the tectum and that the predicted pattern of terminal movement is visible on the tecta of perciform fish.