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PubMed1994

Inhibition of retinal growth cone activity by specific metalloproteinase inhibitors in vitro.

The developing neural retina expresses a set of extracellular proteases including plasminogen activator and gelatinases. Since neurites of retina cells cultured on fluorescent gelatin digest the substrate in their paths, we have suggested that the proteases are used by the tips of growing fibers to allow them to migrate within the mass of the tissue in vivo. In order to obtain further information about relationships between extracellular proteases and fiber growth, we have examined the effects of the specific inhibitors HS-LFA (HS-Leu-Phenylala-Ala, enantiomeric forms 1 and 2), bathophenanthroline sulfonate (BPS), phenylmethyl sulfonyl fluoride (PMSF), and relevant controls on the activity of retinal growth cones in vitro, monitored by time lapse video microscopy. Of the inhibitors tested, only the two enantiomeric forms of HS-LFA caused a reproducible cessation of both spike extension and filopodial processes at the growth cone ruffling, while control media had no effect. In some cases, the growth cone swelled and exhibited small protrusions. The behavior of growth cones was in sharp distinction to that of the cytoplasm of neural cells, and membrane ruffling of flat cells, which continued in activity throughout. Growth cone activity returned after several hours in the presence of the agent. BPS was toxic at concentrations above 2.5 mM. Below that, it had no effect. L-cysteine, PMSF, and control media had no effect. The relevance of these results to the possible role of proteases in fiber outgrowth from retinal cells is discussed.

Animals↗
PubMed2002

Hypothermia-induced platelet aggregation in heparinized flowing human blood: identification of a high responder subpopulation.

Cold-induced platelet aggregation (CIPA) in PRP has previously been documented in connection with platelet preservation (4-15 degrees C). This report describes hypothermia-induced platelet aggregation (HIPA) in whole blood and at temperatures used in open-heart surgery (24-32 degrees C). HIPA (specifically, the formation of occlusive aggregates) was studied in human whole blood. Fresh heparinized (1.5 U/ml) human blood was cooled and maintained at target temperatures (15, 20, 24, 28, 32, or 37 degrees C) as it flowed (1 ml/min) through 75-cm long 1/32 inches internal diameter polymer conduit. The formation of aggregates in the tubing was verified using optical video microscopy and was quantified by a light-scattering method and a constant-pressure filtration method. Donors were tested at least twice at each target temperature and were classified into three separate response regimes (Low, Medium, and High) on the basis of the number of aggregates and the duration of their appearance. The screening of 121 donors (average age 22.3 +/- 4.3 years) for HIPA at 24 degrees C (the temperature of maximum response) indicated 14% High Responders, 18% Medium Responders, and 68% Low Responders. HIPA was inhibited by EDTA, citrate, PGE1, and Tirofiban, but not by aspirin, and it was enhanced by elevated heparin levels. HIPA was consistently noted in the blood of a subpopulation of donors, and the associated platelet aggregates in the blood of High Responders were rigid and occlusive. It is postulated that such aggregates may contribute to cognitive dysfunction noted in patients undergoing hypothermic open-heart surgery, and that postulus is being investigated.

Alprostadil↗
PubMed1995

Chromosomes take an active role in spindle assembly.

The assembly of a bipolar spindle is essential for the accurate segregation of replicated chromosomes during cell division. Do chromosomes rely solely on other cellular components to regulate the assembly of the bipolar spindle or are they masters of their own fate? In the Zhang and Nicklas(1) study reviewed here, micromanipulation techniques and video microscopy were used to demonstrate the different roles that chromosome arms, kinetochores and centrosomes play in bipolar spindle assembly.

Animals↗
PubMed2003

Fluid shear contributions to bacteria cell detachment initiated by a monoclonal antibody.

Receptor-mediated adhesion of bacteria to biological surfaces is a significant step leading to infection. Due to an increase in bacterial antibiotic resistance, novel methods to block and disrupt these specific interactions have gained considerable interest as possible therapeutic strategies. Recently, several monoclonal antibodies specific for the Staphylococcus aureus collagen receptor demonstrated specialized ability to displace attached cells from collagen in static assays. In this study, we experimentally examine the monoclonal antibody detachment functionality under physiological shear conditions to evaluate the role of this parameter in the detachment process. The detachment of staphylococci from collagen was quantified in real-time using a parallel plate flow chamber, phase contrast video-microscopy and digital image processing. The results demonstrate a unimodal dependence of detachment on fluid wall shear rate. The observed decrease in effective detachment rate with increasing force at the highest shear levels evaluated is counterintuitive and has not been previously demonstrated. Several possible mechanisms of this result are discussed.

Antibodies, Monoclonal↗
PubMed2005

Reversible permeabilization using high-intensity femtosecond laser pulses: applications to biopreservation.

Non-invasive manipulation of live cells is important for cell-based therapeutics. Herein we report on the uniqueness of using high-intensity femtosecond laser pulses for reversibly permeabilizing mammalian cells for biopreservation applications. When mammalian cells were suspended in a impermeable hyperosmotic cryoprotectant sucrose solution, femtosecond laser pulses were used to transiently permeabilize cells for cytoplasmic solute uptake. The kinetics of cells exposed to 0.2, 0.3, 0.4, and 0.5 M sucrose, following permeabilization, were measured using video microscopy, and post-permeabilization survival was determined by a dual fluorescence membrane integrity assay. Using appropriate laser parameters, we observed the highest cell survival for 0.2 M sucrose solution (>90%), with a progressive decline in cell survival towards higher concentrations. Using diffusion equations describing the transport of solutes, the intracellular osmolarity at the inner surface of the membrane (x = 10 nm) and to a diffusive length of x = 10 microm was estimated, and a high loading efficiency (>98% for x = 10 nm and >70% for x = 10 microm) was calculated for cells suspended in 0.2 M sucrose. This is the first report of using femtosecond laser pulses for permeabilizing cells in the presence of cryoprotectants for biopreservation applications.

Animals↗
PubMed2003

Actin-based centripetal flow: phosphatase inhibition by calyculin-A alters flow pattern, actin organization, and actomyosin distribution.

Previous studies have suggested that the actin-based centripetal flow process in sea urchin coelomocytes is the result of a two-part mechanism, actin polymerization at the cell edge coupled with actomyosin contraction at the cell center. In the present study, we have extended the testing of this two-part model by attempting to stimulate actomyosin contraction via treatment of coelomocytes with the phosphatase inhibitor Calyculin A (CalyA). The effects of this drug were studied using digitally-enhanced video microscopy of living cells combined with immunofluorescent localization and scanning electron microscopy. Under the influence of CalyA, the coelomocyte actin cytoskeleton undergoes a radical reorganization from a dense network to one displaying an array of tangential arcs and radial rivulets in which actin and the Arp2/3 complex concentrate. In addition, the structure and dynamics of the cell center are transformed due to the accumulation of actin and membrane in this region and the constriction of the central actomyosin ring. Physiological evidence of an increase in actomyosin-based contractility following CalyA treatment was demonstrated in experiments in which cells generated tears in their cell centers in response to the drug. Western blotting and immunofluorescent localization with antibodies against the phosphorylated form of the myosin regulatory light chain (MRLC) suggested that the demonstrated constriction of actomyosin distribution was the result of CalyA-induced phosphorylation of MRLC. Overall, the results suggest that there is significant cross talk between the two underlying mechanisms of actin polymerization and actomyosin contraction, and indicate that changes in actomyosin tension may be translated into alterations in the structural organization of the actin cytoskeleton.

Actins↗
PubMed2004

Motility of a biflagellate sperm: waveform analysis and cyclic nucleotide activation.

The sperm of the freshwater clam Corbicula fluminea are unusual in that they have two flagella, both of which are capable of beating. When Corbicula sperm are removed from the gonad and placed into freshwater, most remain immotile. Video microscopy was used to assess signaling molecules capable of activating Corbicula sperm motility. Experiments using the cAMP analogs dbcAMP or 8-Br-cAMP show that elevating cAMP activates flagellar motility. Treatments with 8-Br-cGMP activated motility in similar numbers of sperm. Treatments with the selective cAMP-dependent protein kinase (PKA) inhibitor H-89 block activation by 8-Br-cAMP but not by 8-Br-cGMP. Similar treatments with the cGMP-dependent protein kinase (PKG) inhibitor Rp-8-pCPT-cGMPS block activation by 8-Br-cGMP but not by 8-Br-cAMP. These results suggest that cAMP and cGMP each work through their specific kinase to activate flagellar motility. Analysis of spontaneously activated freely swimming sperm shows that the two flagella beat with different parameters. The A flagellum beats with a shorter wavelength and a higher frequency than the B flagellum. The observed differences in flagellar waveform indicate that the flagella are differentially controlled.

Animals↗
PubMed1984

Induction of shape transformation in sea urchin coelomocytes by the calcium ionophore A23187.

We have investigated the ability of the Ca++ ionophore A23187 to induce the transformation of petaloid sea urchin coleomocytes to the filopodial form. The response of individual cells to different media was observed with time-lapse phase-contrast video microscopy. In the presence of 1 mM CaCl2, isotonic medium containing 1-5 microM A23187 produces a similar shape transformation to that caused by hypotonic shock. Higher concentrations of ionophore (10-20 microM) induce the formation of filopodia that are thinner and less rigid than those generated by hypotonic shock or low doses of ionophore. A23187 also induces shape transformation in highly flattened cells that do not respond fully to hypotonic shock. The induction of cytoplasmic alkalinization by NH4Cl, methylamine-HCl, or the Na+ ionophore monensin does not induce shape transformation, suggesting that increased intracellular pH is not the stimulus for this process. Ultrastructural changes in cytoskeletal organization were examined in negatively stained detergent-extracted cells. Low doses of ionophore produce filopodia that are indistinguishable from those of hypotonically shocked cells, with actin filament bundles that are straight and cohesive along their entire length. High concentrations of ionophore produce filopodia with filament bundles that branch repeatedly and splay apart near their tips, forming loops and irregular curves. These results suggest that an increase in intracellular free Ca++ concentration acts as the trigger that stimulates coelomocyte shape transformation, but that abnormally high concentrations of intracellular Ca++, produced by high doses of ionophore, interfere with actin filament bundling.

Actins↗
PubMed1984

Nucleotide specificity for reactivation of organelle movements in permeabilized axons.

In a permeabilized axon model, exogenous ATP can reactivate intraaxonal saltatory organelle movements (microscopically visible manifestations of fast axonal transport). We have studied the dependence of the reactivated movements on the ATP concentration and have also examined the nucleotide specificity of the reactivation. Organelle transport was visualized in isolated lobster giant motor axons using Nomarski optics and video microscopy. The axons were permeabilized with saponin, and movement was reactivated with ATP or other nucleotides. Some slight movement was seen with ATP concentrations as low as 10 microM. The velocity and frequency of the reactivated transport increased with increasing ATP concentrations up to about 5 mM. Movement was also reactivated by deoxyadenosine triphosphate, but not by AMP-PNP (a nonhydrolyzable ATP analogue), ADP, or AMP. Although other nucleotides (CTP, GTP, UTP, ITP) could reactivate transport, movement equivalent to that produced by 0.1 mM ATP was only seen with tenfold or greater concentrations of the other nucleotides. This pattern of specificity is consistent with the hypothesis that a dynein-like ATPase, rather than a myosin, is involved in fast axonal transport.

Adenosine Triphosphate↗
PubMed1989

Talin dynamics in living microinjected nonmuscle cells.

To investigate the role of talin in the anchoring of actin-containing stress fibers to the cell membrane of nonmuscle cells, a fluorescent analog of the adhesion plaque protein talin was developed, characterized, and microinjected into living cells. Purified chicken gizzard talin was covalently labeled with the fluorescent dye lissamine rhodamine B sulfonyl chloride. The fluorescently labeled protein was then chromatographed on Sephadex G-25 and DEAE-cellulose in order to remove free dye and denatured protein. The fluorescent talin was able to bind purified vinculin and was localized in adhesion plaques, membrane ruffles, microspikes, and polygonal networks in acetone-permeabilized nonmuscle cells. In cells that were double-stained with fluorescent talin and an affinity-purified anti-talin antibody, a one-to-one correspondence of adhesion plaque staining was seen. Living epithelial cells (PtK2) were microinjected during interphase with fluorescent talin. Computer-enhanced video microscopy was used to document adhesion plaque dynamics such as 1) changes in plaque shape, 2) alterations in plaque positions, and 3) the appearance, growth, and dissolution of plaques. In cells that were followed during mitosis, the adhesion plaques disappeared during cell rounding and then subsequently reappeared upon spreading of the two daughter cells. Treatment of microinjected cells with DMSO in order to disassemble stress fibers resulted in an altered localization of the fluorescent talin. Upon recovery of the cell from the drug, the talin was visualized in its characteristic submembraneous position. These results are the first to document the role and distribution of talin in dynamic processes occurring in living microinjected nonmuscle cells.

Animals↗
PubMed1990

Nucleotide specificity for the bidirectional transport of membrane-bounded organelles in isolated axoplasm.

Video microscopy of isolated axoplasm from the squid giant axon permits correlated quantitative analyses of membrane-bounded organelle transport both in the intact axoplasm and along individual microtubules. As a result, the effects of experimental manipulations on both anterograde and retrograde movements of membrane-bounded organelles can be evaluated under nearly physiological conditions. Since anterograde and retrograde fast axonal transport are similar but distinct cellular processes, a systematic biochemical analysis is important for a further understanding of the molecular mechanisms for each. In this series of experiments, we employed isolated axoplasm of the squid to define the nucleoside triphosphate specificity for bidirectional organelle motility in the axon. Perfusion of axoplasm with 2-20 mM ATP preserved optimal vesicle velocities in both the anterograde and retrograde directions. Organelle velocities decreased to less than 50% of optimal values when the axoplasm was perfused with 10-20 mM UTP, GTP, ITP, or CTP with simultaneous depletion of endogenous ATP with hexokinase. Under the same conditions, TTP and ATP-gamma-S were unable to support significant levels of transport. None of the NTPs tested had a differential effect on anterograde vs. retrograde movement of vesicles. Surprisingly, several inconsistencies were revealed when a comparison was made between these results and nucleoside triphosphate specificities that have been reported for putative organelle motors by using in vitro assays. These data may be used in conjunction with data from well-defined in vitro assays to develop models for the molecular mechanisms of axonal transport.

Animals↗
PubMed1990

Effect of filamin and controlled linear shear on the microheterogeneity of F-actin/gelsolin gels.

We have previously established [Cortese and Frieden, J. Cell Biol. 107:1477-1487, 1988] that actin gels formed under shear are microheterogeneous. In this study, the effect of cross-linking (by chicken gizzard filamin), severing (by plasma gelsolin), and shear on actin microheterogeneity are investigated using fluorescence photobleaching recovery and video microscopy. We find that filamin and shear form microheterogeneous F-actin:gelsolin gels by different mechanisms. Bundling of actin:gelsolin filaments by filamin can be explained by an increase in the apparent length of the filaments due to interfilament binding, resulting in a decrease of the polymer number concentration at which filaments organize into anisotropic phases. Some intrafilament binding of filamin to actin filaments may also be present, and those filaments coated with filamin immobilize more slowly than actin under the same polymerization conditions. The length of F-actin/gelsolin filaments seems to be a major factor in controlling the extent of bundling relative to network formation. In contrast, the effect of shear on the microheterogeneity of actin:gelsolin filaments is consistent with our previous proposal that shear aligns actin filaments, allowing filament-filament interactions and phase formation to occur. Short filaments are unable to organize into branched actin networks, but they can create large aggregates under low shear. Longer actin filaments will exist as networks with variable levels of branching and are less sensitive to shear. The effect of the intensity of a shear field on the spatial distribution of actin may involve a progressively more random orientation of actin molecules and bundles. A regular pattern develops across the sample at low shear rates (0.04-1.39 s-1), and becomes very irregular at higher shear rates (greater than 10 s-1). We suggest here that actin-binding proteins and shear can control the transition between isotropic networks and anisotropic phases by their effect on apparent length and local filament concentration, and also that this transition can have substantial effects on the resistance of cells to mechanical stress.

Actin Cytoskeleton↗
PubMed1991

Cell division in Dictyostelium with special emphasis on actomyosin organization in cytokinesis.

This study focuses on the dynamic reorganization of actin and myosin ("conventional" myosin, or myosin-II) during cytokinesis in D. discoideum. This is the first study identifying the birefringence of the spindle microtubules as well as three sets of microfilamentous structure in Dictyostelium. The change of organization in these fibrillar structures was followed in real-time with video microscopy, using a Universal Polarizing Microscope equipped with polarized-light (POL) and differential interference contrast (DIC) optics combined with digital image processing. High-frequency mitotic cells were obtained by semi-synchronous culture, and high-resolution observations were made by utilizing the agar-overlay method (Yumura et al.: Journal of Cell Biology 99:894-899, 1984). The molecular identity of the birefringent structures was determined by fluorescence microscopy. Through-focus observations were performed with an axial resolution of 0.3 micron depth of field. The actomyosin fibrils show a dramatic reorganization throughout mitosis. The fibrils at the leading lamellipodia disappear, and there is a striking assembly of the cortical actomyosin in pro-metaphase, which is accompanied by a decrease in cell volume. The cortical actomyosin gradually increases through anaphase. After late anaphase, very active polar lamellipodia, with an average life of less than 1 minute, are formed. We confirmed that the polar lamellipodia include actin, but not myosin-II. At the cleavage furrow, the microfilaments form two distinctive structures: circular contractile ring at the equator, and a cortical filament array parallel to the polar axis. Myosin is localized in the contractile ring, but not associated with the axial array of F-actin. Actomyosin in the contractile ring gradually transforms into cortical network at the posterior region of daughter cells. The constriction of the furrow is accompanied by a drastic efflux of water as evidenced by highly active contractile vacuole formation and turbulent motion of minute vesicles connected to the furrow. This study demonstrates the presence of a new microfilament structure, as well as the dynamic property of the contractile ring, and sheds new light on the contractile mechanisms underlying cytokinesis.

Actomyosin↗
PubMed1991

Neural crest cell galvanotaxis: new data and a novel approach to the analysis of both galvanotaxis and chemotaxis.

The galvanotaxis response of neural crest cells that had migrated out of the neural tube of a 56-hr-old quail embryo onto glass coverslips was observed using time-lapse video microscopy. These cells exhibit a track velocity of about 7 microns/min and actively translocate toward the negative pole of an imposed DC electric field. This nonrandom migration could be detected for fields as low as 7 mV/mm (0.4 mV/cell length). We find that this directional migration is independent of the speed of migration and have generated a rather simple mathematical equation that fits these data. We find that the number of cells that translocate at a given angle, phi, with respect to the field is given by the equation N(phi) = exp(a0 + a1cos phi), where a1 is linearly proportional to the electric field strength for fields less than 390 mV/mm with a constant of proportionality equal to KG, the galvanotaxis constant. We show that KG = (150 mV/mm)-1, and at this field strength the cellular response is approximately half maximal. This approach to cellular translocation data analysis is generalizable to other directed movements such as chemotaxis and allows the direct comparison of different types of directed movements This analysis requires that the response of every cell, rather than averages of cellular responses, is reported. Once an equation for N(phi) is derived, several characteristics of the cellular response can be determined. Specifically, we describe 1) the critical field strength (390 mV/mm) below which the cellular response exhibits a simple, linear dependence on field strength (for larger field strengths, an inhibitory constant can be used to fit the data, suggesting that larger field strengths influence a second cellular target that inhibits the first); and 2) the amount of information the cell must obtain in order to generate the observed asymmetry in the translocation distribution (for a field strength of 100 mV/mm, 0.3 bits of information is required).

Animals↗
PubMed1992

Cytokinesis is more rapid in Ha-T24-ras transfected rat embryo fibroblasts than in non-transfected control cells.

It has long been known that neoplastic cells are characterized by increases in cell motility. Earlier studies from this laboratory indicated that mitotic events were also altered in many tumor and experimentally transformed cells and that this included increases in metaphase duration and a reduction in the duration of cytokinesis. The studies presented in this paper were done to determine whether or not transfection of normal rat embryo fibroblasts by the Ha-T24-ras oncogene could also produce such alterations in mitotic events. The results obtained with the use of time lapse video microscopy indicate that neither the duration of metaphase nor the rate of chromosome movement during anaphase was altered but that the rate of furrow progression during cytokinesis occurred at a significantly more rapid rate. Thus, the cellular alterations induced by transfection with Ha-T24-ras accelerate microfilament-dependent cytokinetic furrowing without significant effects on microtubule-dependent mitotic events. One of several possible mechanisms that could account for these observations involves a down regulation of protein kinase C which has been reported to occur in many neoplastic cells including those transformed by ras. Such a hypothesis could also have broader implications because it may be applicable to the increase in motility and metastatic activity generally observed in transformed cells.

Animals↗
PubMed1993

Computerized analysis of tumor cells flowing in a parallel plate chamber to determine their adhesion stabilization lag time.

The importance of cell adhesion in a variety of physiological phenomena requires development of an understanding of the factors and molecular mechanisms underlying these behaviors. Cell adhesion is a multistep process involving primary receptor-ligand interactions followed by secondary events that may lead to the formation of focal contacts. Due to the lack of well-defined assays to study adhesion stabilization, little is known about this process, except that it may involve signaling events, receptor recruitment, and, as we have demonstrated, covalent peptide cross-linking by cell membrane-associated transglutaminase [Menter et al.: Cell Biophys. 18:123-143, 1992). To study the stabilization process we have developed a dynamic assay employing a parallel plate flow chamber coupled with video microscopy and digital image processing. Our studies utilize wheat germ agglutinin-selected human metastatic melanoma cell variants that exhibit differences in their experimental metastatic potential and expression of transglutaminase. Using this assay, quantifying cell-substrate stabilization was found to be quick, reliable, reproducible, and useful in evaluating agents that block this process.

Aged↗
PubMed2003

Some glial progenitors in the neonatal subventricular zone migrate through the corpus callosum to the contralateral cerebral hemisphere.

The great majority of glial cells of the mammalian forebrain are generated in the perinatal period from progenitors in the subventricular zone (SVZ). We investigated the migration of progenitors from the neonatal (postnatal day 0, P0) rat forebrain SVZ by labeling them in vivo with a green fluorescence protein (GFP) retrovirus and monitoring their movements by time-lapse video microscopy in P3 slices. We identified a small number of progenitors that migrated tangentially within the corpus callosum (CC) and crossed the midline. These cells retained a relatively uniform morphology: the leading process was extended toward the contralateral side but showed no process branching or turning away from the migratory direction. Net migration requires the elongation of the leading process and nuclear translocation, and the migrating cells in the CC showed both modes. We confirmed the presence of unmyelinated axon bundles within the P3 CC, but failed to detect any radially directed glial processes (vimentin- or GLAST-immunolabeled fibers) spanning through the CC. Confocal images showed a close proximity between neurofilament-immunolabeled axons and the leading process of the GFP-expressing progenitors in the CC. The destination of the callosal fibers was examined by applying DiI to the right cingulum; the labeled fibers ran throughout the CC and reached the left cingulate and motor areas. The distribution and final fates of the retrovirus-labeled cells were examined in P28 brains. A small proportion of the labeled cells were found in the contralateral hemisphere, where, as oligodendrocytes and astrocytes, they colonized predominantly the cortex and the underlying white matter of the cingulate and secondary motor areas. The distribution pattern appears to coincide well with the projection direction of the callosal fibers. Thus, glial progenitors migrate across the CC, presumably in conjunction with unmyelinated axons, to colonize the contralateral hemisphere.

Animals↗
PubMed1989

Junction between parent and daughter axons in regenerating myelinated nerve: properties of structure and rapid axonal transport.

The primary aim of this work was to investigate the properties of rapid axonal transport in regenerating myelinated axons in the sciatic nerve of Xenopus laevis, with particular attention to events at the junction between the proximal, intact axon (the "parent") and the distal, newly formed axon (the "daughter"). Morphological studies indicated that all myelinated axons initiated regeneration and that at least 80% of these axons regenerated at a rate of 1 mm/day or greater (20 degrees C). The ultrastructure of the junctional region was examined at regeneration times between 3 days and 20 weeks. The main qualitative change in the junctional axoplasm over this period was in its content of particulate organelles. At times up to 2 weeks regeneration, the junction contained abnormal numbers of 50 nm diameter vesicles and 10 nm granules. Between 2 and 5 weeks the junction showed in addition a peripheral rim of large membrane-bounded organelles around a central core of microtubules and neurofilaments. At longer times the numbers of large membrane-bounded organelles diminished and all junctions contained prominent accumulations of 10 nm granules. The rate of rapid axonal transport of protein was similar in parent and daughter axons. Compared to the parent axons, a 2-5 times greater amount of protein was deposited to a stationary phase in daughter axons. Specimens of nerve that were subjected to mechanical stress during the removal of the perineurium showed a large accumulation of rapidly transported protein in the region of the crush at regeneration times up to 40 days; some of the accumulated protein was subsequently transported retrogradely. Video microscopy of isolated axons supplied evidence that the transport deficit in mechanically stressed nerve was a partial block of anterograde vesicle transport, plus a reversal of anterograde transport, at the junction of parent with daughter axons. No structural changes were detected in mechanically stressed nerve. The results show that the junction between parent and daughter myelinated axons is a region with distinct morphology at which the dynamics of anterograde axonal transport may change dramatically.

Animals↗