Browsing by Author "ALVAREZ, J"

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    ANTI-200 KDA NEUROFILAMENT ANTIBODY CROSS-REACTS WITH MICROTUBULE-ASSOCIATED PROTEIN-2 (MAP-2)
    (1989) ALLENDE, ML; KRAUSS, RY; TREMBLAY, C; ALVAREZ, J; INESTROSA, NC
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    ASSEMBLY AND DISASSEMBLY OF AXONAL MICROTUBULES OF THE TOAD XENOPUS-LAEVIS UNDER THE EFFECT OF TEMPERATURE
    (1992) ALVAREZ, J; FADIC, R
    In toads Xenopus laevis living at 11-degrees (winter), the microtubular density of 4-mum myelinated axons of lumbosacral nerves was assessed with the electron microscope. In controls, the density was 11.2 microtubules/mum2. In nerves incubated at 0-degrees, microtubules decreased following a simple exponential curve with a half time of 4.7 min (k = 0.149 min-1); residual microtubules were 4.5%. After rewarming, the full complement of microtubules reappeared within 60 min. In steady state, the microtubular density exhibited a linear relationship with temperature (range: 0-22-degrees; slope 0.94 microtubules/mum2 per degree; r, 0.96). After heating the nerve by 11-degrees above the physiological temperature, microtubules increased by 83%, whereby the pool of unpolymerized tubulin was at least 2.7 mg/ml of axoplasm. A seasonal variation of the microtubular density was observed which accorded with the environmental temperature. The macroscopic kinetics of microtubule disassembly in the axoplasm is similar to that reported for purified tubulin but that of assembly is slower. Microtubules of peripheral axons of Xenopus are cold-labile and vary during the annual cycle.
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    AXONAL MICROTUBULES - COMPARATIVE ANATOMY IN VERTEBRATES, INCLUDING MAN
    (1991) VERGARA, J; SERRA, M; SAITUA, F; ITURRIAGA, R; ALVAREZ, J
    The microtubular density was assessed with the electron microscope in 3-mu-m myelinated fibers, myelin excluded, of 11 species from the following classes: Osteichthyes, Amphibia, Reptilia, Aves, and Mammalia. The average for all species was 20.6 microtubules/mu-m2. Dispersion of values was restricted as shown by a coefficient of variation of 15.8. The microtubular content of nonmedullated axons was assessed in trout, lizard, finch, and man. In the four species, the number of microtubules increased with the cross sectional area of the axon. In trout, lizard and finch, the microtubular density decreased from over 100 microtubules/mu-m2 in fibers smaller than 0.1-mu-m2 to about 30 in 1-mu-m2 fibers; in axons of equal size, the packing of microtubules of nonmedullated was similar between them, and with reported values for peripheral axons of cat and rat. In man, the microtubular density of nonmedullated fibers exhibited only a mild decrease with the axonal size. In the finch, myelinated and nonmedullated axons overlapped in the range 0.23-0.60-mu-m2 and both groups exhibited similar microtubular densities. We conclude that the packing of microtubules of the vertebrate peripheral axon is a feature largely conserved during evolution.
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    AXONAL SPROUTING INDUCED IN THE SCIATIC-NERVE BY THE AMYLOID PRECURSOR PROTEIN (APP) AND OTHER ANTIPROTEASES
    (1992) ALVAREZ, J; MORENO, RD; LLANOS, O; INESTROSA, NC; BRANDAN, E; COLBY, T; ESCH, FS
    Protease inhibition is the mechanism by which some trophic factors promote the extension of neurites. In the rat sciatic nerve, we assessed the ability to induce sprouts of the APP isoform that embodies the Kunitz antiprotease domain and other antiproteases. With the electron microscope, axonal sprouts were found when antiproteases were supplied but not after administration of inactive substances. We conclude that axons have a drive to sprout which can be released by the unbalance of an extracellular protease antiprotease system. We propose that this system is involved in the pathogenesis of Alzheimer's disease.
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    AXONS GROW IN THE AGING RAT BUT FAST TRANSPORT AND ACETYLCHOLINESTERASE CONTENT REMAIN UNCHANGED
    (1988) INESTROSA, NC; ALVAREZ, J
    Caliber and microtubular density of myelinated fibers, acetylcholinesterase (AChE) content and its accumulation at a ligature were studied in the phrenic nerve of mature (3-4 months) and aging (2-year-old) rats. The number of axons remained constant. The cross-sectional area of the nerve was 67% greater in the older group: the axoplasm, though, constituted about 20% of the nerve tissue irrespective of age. The mean cross-sectional area of myelinated axons was twice as big in aging compared to mature rats. All axons grew in the same proportion irrespective of their original caliber. The microtubular density of 3-.mu.m axons was about 22 microtubules .mu.m2 in mature and aging rats. The AChE activity of aging rats was half as much as that of mature rats if it was expressed per wet weight of nerve tissue but did not change if it was expressed per nerve fiber. Twenty-four hours after ligation of the nerve, total AChE activity rose in mature and aging rats by ca. 168%: the molecular forms.sbd.asymmetric and globular.sbd.accumulated in the same proportion in both age groups. We conclude that myelinated axons grow in the adult stage of life but the structure of axoplasm, content of AChE per axon, and rate of fast transport remain lifelong features of nerve fibers.
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    AXONS SPROUT AND MICROTUBULES INCREASE AFTER LOCAL INHIBITION OF RNA-SYNTHESIS, AND MICROTUBULES DECREASE AFTER INHIBITION OF PROTEIN-SYNTHESIS - A MORPHOMETRIC STUDY OF RAT SURAL NERVES
    (1991) BUSTOS, J; VIAL, JD; FAUNDEZ, V; ALVAREZ, J
    Drugs that inhibit RNA or protein synthesis are known to affect some functional properties of axons. In this context, we studied the ultrastructural effects of actinomycin-D, an inhibitor of RNA synthesis, and cycloheximide and emetine, inhibitors of protein synthesis, in rat sural nerves. A silicone sleeve (4 mm long) loaded with drug was placed around the nerves and left for about a week. The ultrastructural alterations of axons and Schwann cells progressed over this period. After cycloheximide and emetine, the cytoplasm of Schwann cells was enlarged and the rough endoplasmic reticulum was prominent. After actinomycin-D, the Schwann cells reached the stage of lysis. Nonmedullated were more affected than myelinated axons. After cycloheximide and emetine, the axoplasmic matrix decreased substantially but reversibly. Microtubules of nonmedullated fibres decreased by about 50%. Actinomycin-D determined sprouting of axons and a rise of axonal microtubules; in nonmedullated axons, the normal inverse correlation between microtubular density and calibre gave way to a high and constant density for all axonal sizes. A few millimetres proximal and distal to the sleeve, the nerve tissue and the axonal microtubular content were close to normal, i.e. the effects of drugs were local. Present results suggest that the local turnover of amino acids in the axon is necessary to maintain the integrity of microtubule and neurofilament proteins. We propose that the Schwann cell down-regulates the axonal cytomatrix.
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    AXOPLASMIC INCORPORATION OF AMINO-ACIDS IN A MYELINATED FIBER EXCEEDS THAT OF ITS SOMA - A AUTORADIOGRAPHIC STUDY
    (1983) ALVAREZ, J; BENECH, CR
    The axoplasmic incorporation of amino acids was studied in the giant Mauthner axon [of Carassius auratus] to determine its magnitude, to estimate the half-life of the resulting material, and to establish whether or not it undergoes transport. The incorporation of lableled leucine, lysine and proline was followed by radioautography. The tracers were injected away from the cell body, either in the floor of the IV ventricle or into the spinal cord. Radioautographic reaction on the Mauthner somata was barely discernible. At the site of injection, for survivals to 5.6 days, a peak of reaction was observed on the Mauthner axoplasm, with a sharp and symmetric decrement in both central and distal directions; the intensity of the peak decreased with survival (14% at 5.6 days). In intact fibers, the intensity of the reaction was 1.4% compared with that over the Nissl substance of neighboring somata; by contrast, in severed fibers whose axoplasm had been directly exposed to the tracers the reaction rose to 4.1%. At early times, the intensity of the response of the axoplasm as a function of survival of the fish did not have a lag time. In high-resolution radioautograms, most of the grains overlay the ground axoplasm. Cycloheximid depressed the reaction over the fiber open to the extracellular space by 55% whereas only by 20% over the intact fiber. Adsorption of free amino acids or their binding to tRNA were ruled out as a cause of the radioautographic response. The results indicated that incorporation of amino acids into macromolecules of the Mauthner fiber occurred locally, and its magnitude over the whole axoplasm was estimted to be one order of magnitude larger than that occurring in the soma. The bulk of the material resulting from this incorporation did nto undergo transport and survived some days. Evidently, the amino acids were incorporated by a ribosomal mechanism.
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    BIOSYNTHESIS OF THE NEUROFILAMENT HEAVY SUBUNIT IN XENOPUS OOCYTES MICROINJECTED WITH RAT-BRAIN POLY(A)+ RNA
    (1987) CROSS, D; ALLENDE, ML; KRAUSS, RY; FUENTES, ME; KALTWASSER, G; ALVAREZ, J; INESTROSA, NC
    In order to study the expression of the major subunit of neurofilaments (NFs), rat brain poly(A)+ RNA was purified by three different procedures and was injected in Xenopus laevis oocytes. This system was able to translate efficiently the 200 kDa NF subunit as shown by a dot-blot immunoassay and by immunoprecipitation of labeled NF polypeptides.
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    BLOOD MARKERS IN ALZHEIMER-DISEASE - SUBNORMAL ACETYLCHOLINESTERASE AND BUTYRYLCHOLINESTERASE IN LYMPHOCYTES AND ERYTHROCYTES
    (ELSEVIER SCIENCE BV, 1994) INESTROSA, NC; ALARCON, R; ARRIAGADA, J; DONOSO, A; ALVAREZ, J; CAMPOS, EO
    In patients with the clinical diagnosis of Alzheimer disease (AD), we searched for systemic changes in components of the blood as a diagnostic tool. The acetylcholine-related enzymes acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) were measured in plasma, erythrocytes, platelets and lymphocytes. Results did not show a general effect; notwithstanding, specific cell types presented alterations either in AChE or BuChE but not in both enzymatic activities. In AD patients, AChE of lymphocytes was reduced by 60% compared with the age-matched controls. However, when patients were divided, the sporadic but not the familial subgroup exhibited a significant reduction. In erythrocytes the BuChE activity was reduced by 45% in sporadic AD. The molecular forms of the lymphocyte AChE were characterized by velocity sedimentation. Both globular forms were subnormal, more so the tetrameric G(4) AChE form than the G(2) form.
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    CALIBERS AND MICROTUBULES OF NERVE-FIBERS - DIFFERENTIAL EFFECT OF UNDERNUTRITION IN DEVELOPING AND ADULT-RATS
    (1990) FAUNDEZ, V; CORDERO, ME; ROSSO, P; ALVAREZ, J
    Sural nerves of 9-week-old rats undernourished since birth, and of adult rats food-restricted for 27 and 48 days, were studied to explore the effect of severe undernutrition on the caliber and microtubules of axons in growing and non-growing animals. In 9-week-old undernourished rats, the number and caliber of myelinated fibers were normal while the cross-sectional area of non-medullated fibers was 29% smaller than controls. By contrast, in adult undernourished rats the cross-sectional area of myelinated fibers was affected sooner and to a greater extent (-28%) than that of non-medullated fibers (-23%). Regardless of age, in both controls and in undernourished rats non-medullatedfibers of equal caliber had similar microtubular content. The same was found in 3-.mu.m myelinated axons. These findings indicate that food restriction affects proportionately caliber and microtubules of axons. It is proposed that the anatomy of the axon is in a dynamic equilibrium and that microtubules participate in the specification of the axonal caliber.
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    DO AXONS GROW DURING ADULTHOOD - A STUDY OF CALIBER AND MICROTUBULES OF SURAL NERVE AXONS IN YOUNG, MATURE, AND AGING RATS
    (1988) SAITUA, F; ALVAREZ, J
    Calibers and microtubules of sural nerve axons were studied in young (6-week-old), mature (14-week-old), and aging (2-year-old) rats.The mean cross-sectional area of nonmedullated fibers was about 0.50 .mu.m2 (range: 0.47-0.52) in the three age groups. Their caliber spectra were also similar. In contrast, myelinated axons grew from 6.6 to 16.7 .mu.m2between the sixth and 14th week of age. The increase of cross-sectinal area was greater, the greater the initial caliber of axon (range 44-154%). No further change of caliber was observed in the aging rat. The cross-sectional area of nerve allotted per myelinated fiber was 42, 66, and 97 .mu.m2 in young, mature, and aging rats, respectively. The fraction of nerve tissue occupied by the axoplasm, though, did not change substantially; it was 20, 28, and 21%, respectively.The microtubular density of 3-.mu.m myelinated axons had a general average of 21 microtubules/.mu.m2. Differences between groups were not significant. In nonmedullated fibers, the microtubular density decreased as the size of the axon increased. No differences were observed between age groups. We conclude that nonmedullated fibers of the sural nerve stop growing before the sixth week whereas myelinated fibers keep growing until the 14th week of age. The correlation between microtubular content and axonal caliber is a lifelong feature of axons.
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    EARLY AXONAL REGENERATION - REPRESSION BY SCHWANN-CELLS AND A PROTEASE
    (1995) TAPIA, M; INESTROSA, NC; ALVAREZ, J
    We have proposed that mature Schwann cells and an extracellular protease repress the sprouting response of axons. To test this hypothesis, we destroyed all cells by freezing a short span of the rat sciatic nerve or inhibited proteases with subperineurial injections of aprotinin, and a crush was made to induce the sprouting response. In unconditioned or in vehicle-injected nerves, axons began to elongate at a constant rate after a delay of about 1 day. The freezing of the nerve distal to the crush obliterated the delay, but the rate of elongation did not change. A similar pattern was observed when the nerve segment was conditioned with aprotinin for 2 days prior to the crush. These effects were abolished when a short untreated segment was left between crush and conditioned region of the nerve. The electron microscopy of the nerve and the immunolocalization of the growth-associated protein (GAP-43) were consistent with the enhanced regrowth observed in conditioned nerves. Our findings support the notion that Schwann cells repress the onset of regeneration and that a local protease is involved. (C) 1995 Academic Press, Inc.
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    INCORPORATION OF AMINO-ACIDS INTO THE AXOPLASM IS ENHANCED BY ELECTRICAL-STIMULATION OF THE FIBER
    (1995) EUGENIN, J; ALVAREZ, J
    The effect of sustained electrical stimulation upon the incorporation of amino acids into the axoplasm was studied in the goldfish Mauthner (M) axon with light autoradiography. An extracellular pulse of tracers applied between M-axons in the medulla resulted in a local and substantial labeling of the M-axoplasm and a faint labeling of the M-perikaryon 4-5 mm away from the site of injection. After 18 h of direct electrical stimulation of the M-axon at 0.3-0.8 Hz, the local incorporation of amino acids into the M-axoplasm doubled. This enhancement declined to reach the baseline within 24 h. A 4 h electrical stimulation did not enhance the incorporation. Transynaptic activation of the M-neuron through the auditory input at 0.1-0.2 Hz for 18 h did not raise the amino acid incorporation in the M-axoplasm. We conclude that electrical discharge of the axon modulates the local incorporation of amino acids into the axoplasm.
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    MICROTUBULAR PACKING VARIES ALONG THE COURSE OF MOTOR AND SENSORY AXONS - POSSIBLE REGULATION OF MICROTUBULES BY ENVIRONMENTAL CUES
    (1989) SAITUA, F; ALVAREZ, J
    In the toad Xenopus laevis, the microtubular density of 3-.mu.m myelinated fibres was assessed in peripheral nerves, dorsal and ventral roots, and dorsal and ventral funiculi of the spinal cord. In the roots, the axonal microtubular density was 6 microtubules/.mu.m2 and twice as much at the other sampling sites. This indicates that the pattern of the microtubular packing may vary along the course of the axon. We propose that axonal microtubules are regulated by local cues.
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    MICROTUBULES AND CALIBERS IN DEVELOPING AXONS
    (1986) FAUNDEZ, V; ALVAREZ, J
    The caliber and microtubular content of growing axons were assessed with the electron microscope in the sural nerve of the rat from birth until the age of 63 days (young adult). The caliber of both nonmedullated and myelinated fibers increased throughout the period of observation. At birth, nonmedullated fibers smaller than 0.2 .mu.m2 represented 69% of the population; at day 63 less than 7% were that small. Irrespective of the age of the rat, the number of microtubule profiles of nonmedullated fibers increased with the cross-sectional area of the axon although their packing decreased. In myelinated fibers of a given caliber, the packing of microtubules increased with time, and by day 63 the density had reached its final value. In nonmedullated fibers of a given caliber, a similar trend was observed after day 31; i.e., fibers showed a small but consistent increase in density. However, before that, and in contrast to myelinated fibers, nonmedullated fibers of defined calibers exhibited a transient increase in the microtubular density. Notwithstanding, during the history of an individual fiber the packing of its microtubules may decrease continuously until stabilizing, because the developing fiber is increasing its caliber and hence decreasing its microtubular density. In the 5-day-old rats, the caliber spectra of myelinated and nonmedullated axons overlapped in the 0.49-0.83 .mu.m2 range and their microtubular densities were similar. We conclude that the microtubular content of developing axons correlates with caliber, an architectural feature, and is largely independent of growth and of its myelinated condition. We propose that the cytoskeletal rather than the transport function commands the organization of axonal microtubules.
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    MICROTUBULES AND CALIBERS IN NORMAL AND REGENERATING AXONS OF THE SURAL NERVE OF THE RAT
    (1986) ESPEJO, F; ALVAREZ, J
    The calibers and microtubular content of axons were studied in normal and regenerating fibers of the sural nerve from 17 to 122 days after a lesion of the sciatic nerve of young adult rats. During this period (70-175 days of age), the cross-sectional area of control myelinated axons almost doubled but that of nonmedullated axons did not change. In regenerating nerves, after 122 days of recovery, the cross-sectional area of myelinated fibers was still 38% below that of the normal side. In contrast, the regenerating nonmedullated population was richer in fine (< 0.2 .mu.m2) and in coarse (> 0.9 .mu.m2) fibers than on the control side; the cross-sectional area averages were 0.50 and 0.54-0.70 .mu.m2 for the normal and regenerating populations, respectively. The microtubular density of normal 3-.mu.m myelinated fibers averaged 24.0 microtubules/.mu.m2. In regenerating fibers of the same size the density varied between 19.2 and 23.2 microtubules/.mu.m2. Microtubular density values of normal and regenerating fibers were not statistically different. In nonmedullated fibers, the microtubular content (expressed as microtubular density or number of microtubules per axon) correlated with the caliber of the fiber. In these correlations, only minor differences were observed between regenerating and uninjured fibers. Our results indicate that nonmedullated fibers terminate their radial growth well before myelinated fibers do, and that axonal microtubular content correlates with the local size of the fiber and is largely insensitive to regeneration.
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