Laser irradiation of centrosomes in newt eosinophils: evidence of centriole role in motility.

Abstract

Newt eosinophils are motile granulated leukocytes that uniquely display a highly visible centrosomal area. Electron microscope and tubulin antibody fluorescence confirms the presence of centrioles, pericentriolar material, and radiating microtubules within this visible area . Actin antibodies intensely stain the advancing cell edges and tail but only weakly stain pseudopods being withdrawn into the cell. Randomly activated eosinophils follow a roughly consistant direction with an average rate of 22 .5 gm/min . The position of the centrosome is always located between the trailing cell nucleus and advancing cell edge . If the cell extends more than one pseudopod, the one closest to or containing the centrosome is always the one in which motility continues .Laser irradiation of the visible centrosomal area resulted in rapid cell rounding . After several minutes following irradiation, most cells flattened and movement continued. However, postirradiation motility was uncoordinated and directionless, and the rate decreased to an average of 14 .5 Am/min . Electron microscopy and tubulin immunofluorescence indicated that an initial disorganization of microtubules resulted from the laser microirradiations . After several minutes, organized microtubules reappeared, but the centrioles appeared increasingly damaged . The irregularities in motility due to irradiation are probably related to the damaged centrioles. The results presented in this paper suggest that the centrosome is an important structure in controlling the rate and direction of newt eosinophil motility .Cell movement is a fundamental process of particular relevance to problems in developmental biology, cell biology, oncology, and immunology. Recent advances in immunofluorescence and electron microscopy have resulted in considerable research on the mechanisms ofcell motility. As a result ofmany studies, it is fairly well accepted that the cytoskeleton is a major element in the cellular motile apparatus (1-5). Cytoskeletal proteins such as actin and myosin are thought to generate forces necessary-for movement (5)(6)(7)(8). In addition to their role in cell division, microtubules (tubulin and associated proteins) have been studied as support structures involved in maintaining cell shape and polarity (5, 9, 10), intracellular partical transport (11), and as structures involved in directed movement of leukocytes (1,12,13) . Recently, the microtubule-organizing centers have been suggested as the structures involved in direction determination (14-17) .

Inhibitory neurons from fetal rat cerebral cortex exert delayed axon formation and active migration in vitro

“…In some types of cells, the centrosome and the Golgi apparatus translocate in the direction of cell migration (Gotlieb et al, 1981;Kupfer et al, 1983;Koonce et al, 1984). For example, relocalization of centrosomes to the site of newly emerging processes has been observed in cultured cerebellar granule neurons (Zmuda and Rivas, 1998).…”

Centrosome positioning in vertebrate development

“…However, whether the centrosome is ahead of or behind the nucleus depends on the cell type (Yvon et al, 2002). For example, the centrosome is located ahead of the nucleus during migration in eosinophils (Koonce et al, 1984) and Chinese hamster ovary (CHO) cells (Yvon et al, 2002), but behind the nucleus in PtK cells in wounded epithelial sheets (Yvon et al, 2002). However, when PtK cells migrate individually instead of in the context of a continuous cell sheet, the position of the centrosome relative to the nucleus does not correlate with the direction of cell migration (Danowski et al, 2001).…”

Mediation of chemoattractant-induced changes in [Ca2+]i and cell shape, polarity, and locomotion by InsP3, DAG, and protein kinase C in newt eosinophils.

“…Newt eosinophils were prepared by a modification of the technique of Koonce et al (1984). Newts were bled by clipping the tip of the tail and 0.2-0.3 ml of blood collected in 1 ml heparinized amphibian culture medium (ACM, pH 7.2, and containing, mM: 64 NaC1, 4.3 KC1, 1.4 CaCI2, 1.3 MgSO4, 0.7 NaH2PO4, 19.2 Hepes and 4.4 glucose).…”