bluuebunny/crossref_metadata_2025_split huggingface dataset:
The antagonistic effects of 5‐hydroxytryptamine and methylxanthine on the gill cilia of mytilus edulis
Authors: Sanderson Michael J. , Dirksen Ellen R. , Satir Peter 
Published: February 2005

AbstractThe laterofrontal (LF) cirri on isolated gill filaments of Mytilus edulis, prepared in natural seawater, are active and initially beat with an average frequency of about 8 Hz (with a range of 6–14 Hz). However, the lateral (L) cilia on these filaments are arrested in a position at the end of their recovery stroke. Perfusion of the filament with artificial seawater (ASW), with or without 1% ethanol, has little or no biological effect on the activity of the LF cirri, although a transitory decrease in frequency often accompanies the perfusion process. The L cilia remain arrested during perfusion with ASW. The exposure of the gill to low levels of 5‐hydroxytryptamine (5HT) (10−8 < 5HT < 10−7 M) has no effect on the activity of the LF cirri but stimulates the L cilia to beat. Exposure to higher concentrations of 5HT (> 10−7 M) elevates the beat frequency of the L cilia and simultaneously inhibits the activity of the LF cirri, leading to their arrest in a position at the end of the effective stroke. This arrest of the LF cirri occurs as the L cilia attain a 5HT‐induced beat frequency between 12 to 14 Hz. The influence of 5HT on the L cilia and the LF cirri can be reversibly mimicked or enhanced by the phosphodiesterase inhibitor 3‐isobutyl‐1‐methylxanthine (IBMX). A concentration of 0.5 mM IBMX mimics low 5HT concentrations (about 10−7 M) by stimulating the L cilia to beat without affecting the beat frequency of the LF cirri. A combimation of 10−7 M 5HT and 0.5 mM IBMX in ASW mimics high (> 10−6 M) 5HT concentrations by arresting the LF cirri and increasing the beat frequency of the L cilia. Under these conditions, the threshold of the LF cirri arrest response is again found to occur as the L cilia attain a beat frequency of 12 – 14Hz. These results suggest that the mechanisms of LF cirri arrest and L cilia activation are mediated by 5HT ‐induced changes in intracellular cyclic AMP levels.
Localization of actin in Chlamydomonas using antiactin and NBD‐phallacidin
Authors: Detmers Patricia A. , Carboni Joan M. , Condeelis John 
Published: February 2005

AbstractWe have localized actin in gametes of Chlamydomonas reinhardi by two approaches: (1) indirect immunofluorescence with an affinity‐purified antibody and (2) staining with NBD‐phallacidin, a fluorescent reagent that binds only to F‐actin [Barak et al, 1980, Proc Natl Acad Sci, 77:980–984]. Staining of either mating type “plus” (mt+) or “minus” (mt−) gametes with antiactin antibody resulted in similar fluorescent images: most of the actin was located peripherally along the lateral and posterior aspects of the cells. There was diffuse staining centrally, but the flagella did not stain. No brightly stained spot was observed near the mt+ mating structure, the site where the fertilization tubule elongates with concomitant polymerization of actin [Detmers et al, 1983, J Cell Biol, 97:522–532]. Gametes stained prior to mating with NBD‐phallacidin showed no fluorescence above background, indicating that there were no concentrations of F‐actin in these cells. This suggested that the cytoplasmic staining observed with antiactin represented primarily a nonfilamentous form of the protein. In mating gametes staining with NBD‐phallacidin was detected only in the fertilization tubule, indicating that this was the only dense accumulation of filamentous actin within the cells. Mating gametes stained with antiactin antibody exhibited cytoplasmic fluorescence that was slightly more punctate than prior to mating, and the fertilization tubule was brightly stained. Our observations suggest that the site‐specific polymerization of actin within the fertilization tubule occurs in the absence of a concentrated pool of actin subjacent to the mating structure.
Alpha actinin distribution and extracellular matrix products during somitogenesis and neurulation in the chick embryo
Authors: Lash James W. , Ostrovsky David , Mittal Balraj , Sanger Joseph W. 
Published: February 2005

AbstractA discrete stage in two different morphogenetic processes has been examined employing fluorescently labelled alpha‐actinin as a probe to localize native alpha‐actinin and antibodies to localize fibronectin and collagen type I. The stage of somitogenesis examined is the transition from the compact mesenchymal somitic mass to the epithelial somitic vesicle (ie, epithelialization of the somite). The stage of neurulation examined is the transition from the relatively flat neuroepithelium to the approximation of the neural folds. Before these morphogenetic movements begin, the neuroepithelium is sitting upon a basal lamina and interstitial collagen, and the somite is surrounded by a meshwork of interstitial collagen. During both of these processes, the cells become narrowed at their apices in the region of the tissue that is becoming concave, and alpha‐actinin is localized in the apices. The localization of intracellular alpha‐actinin and extracellular fibronectin, and the distribution of collagen, suggest that there is a coordinated appearance and distribution of these molecules that is temporally associated with these discrete morphogenetic events.
Regulation of sperm flagellar movement by protein phosphorylation and dephosphorylation
Authors: Murofushi Hiromu , Ishiguro Koichi , Takahashi Daisuke , Ikeda Jun , Sakai Hikoichi 
Published: February 2005

AbstractFlagellar motility of Triton models of sea urchin spermatozoa was reactivated by cyclic AMP‐dependent protein kinase and a protein factor, termed motility activator, both of which were prepared from the detergent‐extract of sea urchin spermatozoa. It was shown that phosphorylation of the motility activator by the protein kinase is necessary for the reactivation of flagellar motility [Ishiguro et al, J. Cell Biol. 92:777–782, 1982; Murofushi et al, in “Biological Functions of Microtubules and Related Structures,” Academic Press, 1982]. Reactivating factor was also detected in a KCI‐extract of the axoneme fraction devoid of the detergent‐extractable materials. The activity of this factor was also cyclic AMP‐ and protein kinase‐dependent. Furthermore, when freshly prepared Triton models were treated with phosphoprotein phosphatase prepared from bovine cardiac muscle, the flagellar motility was drastically suppressed. This inhibition of the motility was partially recovered by the addition of cyclic AMP and protein kinase to the phosphatase‐treated models.
Incorporation and turnover of labeled exogenous tubulin in the mitotic spindles of Chaetopterus oocytes and HeLa cells
Authors: Goode Dennis , Sarma Vidya 
Published: February 2005

AbstractThe incorporation of tubulin into mitotic spindles in situ was studied by incubating permeabilized mitotic cells in solutions containing [3H]GTP‐labeled or dichlorotriazinylamino fluorescein (DTAF)‐labeled tubulin. Metaphase HeLa cells or spindle‐containing “minicells” from Chaetopterus oocytes were lysed in a microtubule‐assembly buffer plus 0.5% Nonidet P‐40, 1 mg/ml 120,000g supernatant mammalian brain tubulin, and [3H]GTP. After different periods of incubation, mitotic spindles were isolated in 2 M‐glycerol‐containing assembly buffer and separated from unbound counts by centrifugation through a 4 M‐glycerol cushion; 3H counts per mg protein increase linearly for 8–12 min and then reach a plateau or steady state in both Chaetopterus oocytes and HeLa cells. Addition of 4 mM CaCl2 blocks or reverses incorporation. Little or no [3H]GTP is incorporated if exogenous tubulin or lysed cells are omitted from the assembly mixture.To measure the loss rate of [3H]GTP‐tubulin from mitotic spindles, cells were incubated in tubulin plus [3H]GTP for 30 min, and a 20‐fold excess of cold GTP (2 mM) was added. Samples were removed after incubation for different periods, and spindles were isolated as described above and counted for 3H content. [3H]GTP is lost from spindles at a rate of about 16%/min until a new steady state is reached in about 8 min. These results are consistent with an incorporation and turnover of [3H]GTP‐tubulin in spindle microtubules of these lysed‐cell models.The location of this newly incorporated tubulin in the spindle was investigated by incorporating fluorescent DTAF‐tubulin into mitotic spindles of these lysed cell types. A short pulse (2–5 min) appears to label microtubules (MTs) near metaphase chromosomes and longer exposures label the entire spindle.The rates of incorporation and turnover that we see by [3H]GTP and fluorescent tubulin incorporation in situ are faster than those observed with brain MTs at steady state in vitro but are in the range of the rates of spindle fiber formation in prophase, and spindle MT reassembly after cooling.
Motility and centrosomal organization during sea urchin and mouse fertilization
Authors: Schatten Heide , Schatten Gerald 
Published: February 2005

AbstractMotility and the behavior and inheritance of centrosomes are investigated during mouse and sea urchin fertilization. Sperm incorporation in sea urchins requires microfilament activity in both sperm and eggs as tested with Latrunculin A, a novel inhibitor of microfilament assembly. In contrast the mouse spermhead is incorporated in the presence of microfilament inhibitors indicating an absence of microfilament activity at this stage. Pronuclear apposition is arrested by microfilament inhibitors in fertilized mouse oocytes. The migrations of the sperm and egg nuclei during sea urchin fertilization are dependent on microtubules organized into a radial monastral array, the sperm aster. Microtubule activity is also required during pronuclear apposition in the mouse egg, but they are organized by numerous egg cytoplasmic sites. By the use of an autoimmune antibody to centrosomal material, centrosomes are detected in sea urchin sperm but not in unfertilized eggs. The sea urchin centrosome expands and duplicates during first interphase and condenses to form the mitotic poles during division. Remarkably mouse sperm do not appear to have the centrosomal antigen and instead centrosomes are found in the unfertilized oocyte. These results indicate that both microfilaments and microtubules are required for the successful completion of fertilization in both sea urchins and mice, but at different stages. Furthermore they demonstrate that centrosomes are contributed by the sperm during sea urchin fertilization, but they might be maternally inherited in mammals.
Cell shape and organization of F‐actin and microtubules in randomly moving and stationary amebae of Dictyostelium discoideum
Authors: Roos U.‐P. , De Brabander M. , Nuydens R. 
Published: February 2005

AbstractWe investigated, by video‐light microscopy and fluorescence microscopy with probes specific for microtribules (MTs) and F‐actin, the relationship between cytoskeletal elements, cell shape and behavior of vegetative, undifferentiated amebae of Dictyostelium discoideum, strain NC‐4. In an unconstrained situation, as on the underside of a coverglass in a thin layer of liquid medium, interphase cells moved around randomly in a polypodial or monopodial fashion. Locomotion was characterized by the formation of pseudodigits, rounded or pointed pseudopodia, and retraction fibers. F‐actin occurred in all these structures, as well as in a thin cortical layer. Microtubules extended into some of the cellular extensions rich in F‐actin. Pseudopodial activity, but not locomotion, also took place at the interface between medium and air, demonstrating that ameboid movement requires contact with a solid substrate. Stationary mitotic amebae on glass were studded with continuously changing, peripheral spike‐shaped filopodia that also contained F‐actin. During telophase and cytokinesis the spikes were gradually replaced by pseudopodia in transition to the fully motile phase.In live cells, the nucleus‐associated body (NAB), which is at the center of the complex of cytoplasmic MTs [CMTC; term from Brinkley, Fuller, and Highfield, 1975] was in a rather fixed position; it did not orient in a concerted fashion to follow changes in the direction of cell movement. In amebae fixed and processed for fluorescence microscopy after a period of recorded movement, the NAB was not preferentially positioned with respect to the nucleus and the direction of movement. It is unlikely that the NAB exerts a directional control during randon ameboid movement. The complex of cytoskeletal MTs must be very dynamic or flexible to adjust to the rapid changes of cell shape.
Rearrangement of tubulin, actin, and myosin in cultured ventricular cardiomyocytes of the adult rat
Authors: Guo Juan‐Xia , Jacobson Stuart L. , Brown David L. 
Published: February 2005

AbstractAntitubulin, phalloidin. and antimyosin were used to study the distribution of microtubules, microfilaments, and myofibrils in cultured adult cardiomyocytes. These cells undergo a stereotypic sequence of morphological change in which myotypic features are lost and then reconstructed during a period of polymorphic growth. Microtubules, though rearranged during these events in culture, are always present in an organized network. Myosin and actin structures, on the other hand, initially degenerate. This initial degeneration is reversed when a cell attaches to the culture substratum. Upon attachment, new microtubules are laid down as a cortical network adjacent to the sarcolemma and, subsequently, as a network in the basal part of the cell. Actin and then myosin filament bundles appear next, in a pattern corresponding to the pattern of the microtubules. Finally, striated myofibrils are formed, first in the central part of the cell, and subsequently in the outgrowing processes of the cell, A mechanism is suggested by which the eventual polymorphic shape of a cell is related to the shape of its initial area of contact with the culture substratum. Finally, a model of myofibrillogenesis is proposed in which microtubules participate in the insertion of myosin among previously formed actin filament bundles to produce myofibrils.
Naturally occurring tubulin‐containing paracrystals in Allogromia: Immunocytochemical identification and functional significance
Authors: Rupp Gerald , Bowser Samuel S. , Mannella Carmen A. , Rieder Conly L. 
Published: February 2005

AbstractBundles of microtubules (MTs) are readily visualized in vivo by videomicroscopy in highly flattened reticulopodia of the foraminiferan protozoan Allogromia sp. strain NF. In this report we use videomicroscopy, immunocytochemistry, and high‐voltage electron microscopy to characterize the dynamic changes that occur in this extensive MT cytoskeleton, and in the associated cytoplasmic transport, during induced withdrawal and subsequent reextension of reticulopodia. Within seconds after application of the withdrawal stimulus (seawater substitute made hypertonic with MgCl2) intracellular bidirectional transport along linear MT‐containing fibrils ceases and is replaced by an inward, constant‐velocity flow of cytoplasm along the fibrils. As withdrawal continues, most fibrils become wavy and coalesce to form phase‐dense pools. These wavy fibrils and phase‐dense pools contain a paracrystalline material and few if any MTs. Same‐section correlative immunofluorescence and high‐voltage electron microscopy reveal that the paracrystalline material contains tubulin. During recovery linear fibrils (MTs) rapidly extend from the phase‐dense pools (paracrystals), which concurrently shrink in size, thus reestablishing normal network morphology and motility. We conclude that the MT cytoskeleton in Allogromia reticulopodia is transfonned during withdrawal into a tubulin‐containing paracrystal, which serves as a temporary reservoir of MT protein and an initiation site for MT regrowth.
Relationship between intermediate filaments and microfilaments in cultured fibroblasts: Evidence for common foci during cell spreading
Authors: Green Kathleen J. , Talian John C. , Goldman Robert D. 
Published: February 2005

AbstractSpreading and fully spread chick embryo fibroblasts (CEF) were examined by double‐label fluorescence microscopy using the actin‐specific probe rhodamine‐phalloidin and an antibody directed against CEF intermediate filaments (IF). During midspreading, a striking relationship became discernible: statistical analysis showed that approximately half of the cell population exhibited one or more phase‐dense, phalloidin‐binding nodules that appeared to act as foci from which IF diverged. Coincidence between actin‐containing structures and IF was not limited to these centers; IF could also frequently be seen running in close parallel arrays with stress fibers.Ultrastructural analysis confirmed the presence of non‐membrane‐bound out‐pocketings along the length of stress fibers from which 10‐nm IF diverged. These structures varied in size and shape, and displayed a dense, fine fibrillar appearance. IF and microfilaments (MF) were distinguished by size and by decoration of MF with myosin subfragment‐1. Other IF‐MF interactions were seen in cells of all stages: IF were observed to loop through stress fibers, most frequently at the cell margins. In colchicine‐treated cells, IF became redistributed into cables that often ran parallel and appeared to merge with stress fibers. Cytochalasin D‐treated CEF exhibited loose aggregates of actin‐containing material that appeared to be associated with IF.These results suggest the possibility of an interaction between actin‐containing structures and IF, particularly during cell spreading in cultured fibroblasts.
Effects of pteridines on the filopodia of Dictyotelium discoideum vegetative amoebae
Authors: Rifkin Jared L. , Wali Abdul W. 
Published: February 2005

AbstractLiving vegetative amoebae of NC‐4H Dictyostelium discoideum were studied to determine if a variety of pteridines had any effect on the filopodia. We observed that production, elongation, and branching of these filopodia were stimulated by pteridines that are chemoattractants for cells of this strain. This stimulation occurs at chemotactically effective concentrations and is observed before motility is evident. A relationship between filopodia and chemoattractant signal processing is discussed.
Bivalent orientation and behavior in crane‐fly spermatocytes recovering from cold exposure
Authors: Janicke Marie A. , LaFountain James R. 
Published: February 2005

AbstractAt metaphase in crane‐fly primary spermatocytes, the two sister kinetochores at the centromere of each homologue in a bivalent normally are adjacent and face the same pole; one homologue has all its kinetochore microtubules (kMTs) extending toward one pole and its partner has all its kMTs extending toward the opposite pole. In contrast, during recovery from exposure to 2°C, one or both homologues in many metaphase bivalents had bipolar malorientations: all kMTs of one kinetochore extended toward one pole and some or all those of its sister extended toward the other. Metaphase sister kinetochores that had most of their kMTs extending toward the same pole were adjacent, and those with most extending toward opposite poles were separated from each other. Distances between homologous centromeres were similar to those in properly oriented bivalents. Maloriented bivalents were tilted relative to the spindle axis, and analysis of living cells showed that tilted configurations were rare during prometaphase in untreated cells but frequently arose in cold‐recovering cells as initial configurations, then persisted through metaphase. This was in contrast to unipolar configurations of bivalents (configurations suggesting orientation of both homologous centromeres toward the same pole), which always reoriented shortly after the configuration arose. We conclude that in cold‐recovering cells, bipolar malorientations are more stable than unipolar malorientations, and the orientation process is affected such that bipolar malorientations arise in bivalents upon initial interaction with the spindle and persist through metaphase.
Alpha‐actinin accumulation in the cortex of echinoderm eggs during fertilization
Authors: Hamaguchi Yukihisa , Mabuchi Issei 
Published: February 2005

AbstractRedistribution of alpha‐actinin during fertilization was investigated by means of the microinjection of fluorescently labeled egg alpha‐actinin in the sea urchin, Hemicentrotus pulcherrimus. Upon fertilization, labeled alpha‐actinin accumulated locally around the sperm binding site, where the fertilization cone formed soon afterwards. The accumulation propagated all over the cortex within 10 sec after a latent period of 10–20 sec. When an egg in Na‐free seawater was injected with both alpha‐actinin and calcium buffer (intracellular free Ca2+ concentration = 9 μM), the accumulation of alpha‐actinin was similar to that in normal seawater, which suggests that the accumulation did not depend on the increase in intracellular pH but only on the increase in the intracellular free Ca2+ concentration. In immature oocytes the accumulation was detected in the cortical region, including the huge protruding cytoplasm where the sperm entered. When labeled egg alpha‐actinin was injected into starfish (Asterias amurensis) oocytes followed by insemination, it accumulated in the cortical layer in a manner similar to the case of sea urchin, except that the accumulation in fertilization cones of maturing oocytcs or reception cones of immature oocytes appeared ringlike and rodlike, respectively. Moreover, just after the arrival of the meiotic apparatus, egg alpha‐actinin accumulated in the cortical region, where the formation of the polar body was expected. This suggests that the meiotic apparatus somehow induced the differentiation of the cortex so as to form a polar body. It is concluded that the cortical region where alpha‐actinin accumulated coincided with the microfilament‐rich region. This suggests that alpha‐actinin plays a role in forming the cortical meshwork of actin filaments.
Is the hemidesmosome a half desmosome? An immunological comparison of mammalian desmosomes and hemidesmosomes
Authors: Jones Jonathan C. R. , Yokoo Karen M. , Goldman R. D. 
Published: February 2005

AbstractAlthough the mammalian epidermal basal cell hemidesmosome bears some superficial resemblance to one half of a desmosome at the ultrastructural level, examination of the structure of the electron‐dense submembranous plaques of the hemidesmosome and desmosome reveals that they differ with respect to their overall morphology and dimensions. Based on these findings, we wondered whether components of the desmosome are present in the hemidesmosome. In order to determine this we prepared a number of stratified squamous epithelial tissues for indirect immunofluorescence using antibody preparations directed against known desmosome components including desmoplakin and certain glycoproteins. These antibody preparations do not show reaction with hemidesmosomes by indirect immunofluorescence criteria. We have also utilized bullous pemphigoid (BP) autoantibodies that have been shown to recognize hemidesmosomes in mammalian skin cells [Mutasim et al., J. Invest. Derm., 84:47–53, 1985]. Double label indirect immunofluorescence observations of neonatal mouse skin prepared using desmoplakin antibodies and BP autoantibodies reveal that hemidesmosomes that are stained by the BP autoantibodies are not recognized by the desmoplakin antibodies. We confirmed these findings at the ultrastructural level by indirect immunogold localization of desmoplakin antibodies and BP autoantibodies. Therefore, the hemidesmosome does not appear to be one half of a desmosome and may possess a very different molecular organization relative to the desmosome. We raise the possibility that the variability between the hemidesmosome and desmosome that we detect at the morphological and immunological level may reflect the functional differences of these two types of junctions.