摘要:

AbstractThe effects of basic fibroblast growth factor (bFGF) on myelin basic protein (MBP) gene expression and myelin‐like membrane formation were investigated in oligodendrocyte cultures containing mainly mature oligodendrocytes expressing MBP. These cultures were obtained by selective detachment of the cells of the oligodendrocyte lineage from 40‐day‐old mixed cultures derived from newborn rat brain. They were further purified by a 3‐day pretreatment with cytosine arabinoside (ARA‐C) in order to kill cycling cells. After withdrawal of ARA‐C, daily treatment of the cells with bFGF for 3 days induced a drastic decrease in MBP mRNA level compared to control cultures treated only with ARA‐C. Moreover, the percentage of oligodendrocytes labelled with anti‐MBP antibodies decreased by 50%, as well as that of oligodendrocytes expressing myelin oligodendrocyte glycoprotein (MOG), whereas proteolipid protein (PLP) immunolabelled cells were less affected. At the ultrastructural level, myelin‐like membranes were still abundant in the ARA‐C‐and bFGF‐treated cultures, but they were conspicuously uncompacted compared to cultures only pretreated with ARA‐C. These results bring the first evidence that bFGF is able to down‐regulate myelin protein gene expression in mature oligodendrocytes and to alter myelin structure. They imply that if bFGF is secreted after a demyelinating lesion of the central nervous system (CNS), this plasticity of mature oligodendrocytes will allow final remyelination of axons to complete only after this factor has returned to low lev

ISSN:0360-4012    

DOI:10.1002/jnr.490400302

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractNeuronal growth cones move actively, accompanying changes in intracellular Ca2+concentration. The movement of growth cones may partly depend on the actomyosin system, considering the presence of actin and myosin II. Yet, Ca2+‐sensitive regulatory proteins for the actomyosin system have not been identified in growth cones. In the present study, caldesmon, an inhibitory protein on actin‐myosin interaction, was detected in the growth cone fraction isolated from embryonic rat brain, using immunoblotting with the antibody to chicken gizzard caldesmon. Morphological evidence of caldesmon in growth cones of cultured rat neurons was obtained using the indirect immunofluorescence method. Since inhibition of caldesmon on actin‐myosin interaction can be overcome by calmodulin and Ca2+, caldesmon may be involved in the Ca2+‐dependent regulation in growth cone motility. Tropomyosin is another member of the actomyosin system whose function may be regulated by caldesmon in smooth and nonmuscle cells. A low Mr isoform of tropomyosin was distributed in the growth cone fraction. Using specific antibodies against tropomyosin isoforms, we further clarified morphologically that the low Mr isoform was localized in growth cones, but not the high Mr isoform. High Mr isoforms of tropomyosin were present in nonneuronal cells. Actin filaments in growth cones may be unstable, since low Mr tropomyosin binds to actin filaments with a lower affinity than high Mr isoforms. The instability of actin filaments may be suitable for the rapid movement and shape changes of growth cones. © 1995 Wiley

ISSN:0360-4012    

DOI:10.1002/jnr.490400303

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractDuring the perinatal period, oligodendroglial precursor cells proliferate rapidly, then cease dividing and differentiate into oligodendroglia. Many of these newly formed oligodendroglia are destined to die. We now demonstrate that oligodendroglia generated in passaged cultures of rat forebrain oligodendroglial precursor cells after removal of basic fibroblast growth factor (basic FGF) from the medium often undergo internucleosomal DNA nicking and nuclear fragmentation, features characteristic of apoptosis. These alterations are rare in cultures maintained continuously in basic FGF. As in many other cellular lineages susceptible to apoptosis, these degenerative changes can be prevented by treatment with the endonuclease antagonist, aurintricarboxylic acid, or by inhibiting de novo RNA or protein synthesis. Supplementation of the basic FGF‐free medium with insulin, insulin‐like growth factor‐1, platelet‐derived growth factor, or ciliary neuronotrophic growth factor also diminishes DNA nicking. Both oligodendroglial differentiation and DNA nicking are induced in basic FGF‐treated cultures by inhibiting DNA synthesis with aphidicholin or excess thymidine, thus suggesting a close linkage between the anti‐apoptotic, anti‐differentiation, and mitogenic effects of basic FGF on the oligodendroglial lineage. © 1995 W

ISSN:0360-4012    

DOI:10.1002/jnr.490400304

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThe use of non‐neural conduits to bridge gaps in peripheral nerves has been noted in the literature for many years. A logical extension of this concept is the introduction of neurotrophic or growth promoting factors into the lumen. We present here an injectable nerve guide that allows percutaneous access to the microenvironment of the regenerating peripheral nerve within the guide's lumen. Hyaluronic acid, a compound associated with decreased scarring and improved fibrin matrix formation, is added sequentially to the regenerating peripheral rat sciatic nerve via this injectable nerve guide. Assessment of nerve regeneration and reinnervation shows better conduction velocity, higher axon counts, and a trend toward earlier myelination with hyaluronic acid compared with saline. This work not only implies hyaluronic acid's role as an agent that aids nerve growth but also describes a new tool that allows percutaneous access to the milieu of a regenerating nerve. © 1995 Wiley‐Liss,

ISSN:0360-4012    

DOI:10.1002/jnr.490400305

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThe proliferative capacity of oligodendrocytes was investigated in the spinal white matter of the twitcher mouse, a murine model of a genetic demyelinating disease globoid cell leukodystrophy (GLD), in which degeneration of oligodendrocytes due to metabolic perturbation has been well documented. In normal mice at 30 and 45 days of age, proliferating cells labeled with 5‐bromo‐2′‐deoxyuridine (BrdU) were scarce, and the majority of BrdU‐labeled cells did not immunostain with antibodies for oligodendrocytes, astrocytes, or microglia/macrophages. Only a few cells with markers for oligodendrocytes, carbonic anhydrase (CA), or the Pi form of glutathione‐S‐transferase (Pi), were labeled with BrdU. In the twitcher spinal cord, total numbers of BrdU‐labeled cells were almost 6 times that of the normal littermate mice at 30 days of age, and 28 times at 45 days of age. However, this increase was largely due to an increase of cells labeled with F4/80, a marker for the microglia/macrophages. CA or Pi positive cells only constituted less than 10% of all labeled cells. With progression of demyelination from 30—45 days, total numbers of CA positive or Pi positive oligodendrocytes decreased, but percentages of cells double‐labeled with BrdU and CA or Pi remained fairly constant. The results indicated that oligodendrocytes proliferated, to some extent, in the twitcher despite the genetic metabolic defect, and their decrease in number with progression of disease was not due to declined proliferation but rather cellular degeneration as the result of an intrinsic metabolic perturbation. ©

ISSN:0360-4012    

DOI:10.1002/jnr.490400306

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractCellular volume and potassium contents were determined in rat astrocytes from primary culture following suspension in isoosmotic (269 mOsm) and hypoosmotic (136 mOsm) phosphate‐buffered saline (PBS) containing various potassium concentrations. Within 1 min of suspension in hypoosmotic PBS, cells swelled to 135% of their volume in isoosmotic PBS. This initial swelling was not altered by varying the potassium concentration of the hypoosmotic PBS. After suspension in hypoosmotic PBS containing 3.2 mM potassium, a regulatory volume decrease (RVD) was observed. Higher concentrations of potassium in hypoosmotic PBS inhibited RVD following osmotic swelling. Cells swollen in hypoosmotic PBS containing 50 mM potassium continued to swell for 7 min, reaching a volume of 141 % of their initial isoosmotic volume. After 7 min, these cells demonstrated a subsequent decrease in volume. The swelling observed between 1–7 min after suspension in hypoosmotic PBS containing 50 mM potassium was not affected by 10 μM gadolinium, 1 mM quinine, 1mM DIDS (4,4′‐diisothiocyanato‐2,2′‐stilbenedisulfonic acid), 1 mM SITS (4‐acetamido‐4′‐isothiocyanato‐2,2′‐stilbenedisulfonic acid), 1 mM furosemide, or 100 μM bumetanide. Normal RVD was obtained in hypoosmotic PBS containing 50 mM potassium, if chloride was replaced with gluconate (but not nitrate) to reduce the extracellular K‐C1 product to that of hypoosmotic PBS containing 3.2 mM potassium. The volume decrease seen between 7–30 min after exposure to hypoosmotic PBS containing 50 mM potassium was blocked by 1 mM DIDS, 1 mM SITS, or 1 mM furosemide. Cellular potassium content was elevated by approximately 60% after 7 min exposure to isoosmotic or hypoosmotic PBS containing 50 mM potassium. In hypoosmotic PBS, this increase in cellular potassium was reduced with replacement of chloride by gluconate, but not by nitrate. The results indicate that astrocytes swollen in PBS containing elevated potassium concentrations continue to swell, in part, by accumulation of potassium plus chloride mediated by an approach to Donnan equilibrium. Cotransport carriers or stretch‐activated channels do not play a role in the enhanced swelling observed in hypoosmotic PBS containing 50 mM potassium. We suggest that a voltage‐sensitive chloride channel mediates this continuation of cell swelling. This mechanism may be important in the persistent swelling of astrocytes observed in pathologic conditions such as trauma and seizures where extracellular potassium is elevated, or when other factors are present which may cause astroglial d

ISSN:0360-4012    

DOI:10.1002/jnr.490400307

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractDuring the early stages of nerve implantation, we followed the dynamic properties of the lateral gastrocnemius muscle of the rat, reinnervated with an acutely or chronically severed peroneal nerve. The aim of this study was to ascertain whether (1) the better functional recovery of a muscle reinnervated by a chronically severed foreign nerve is present from the onset of reinnervation, and (2) whether such functional improvement is due to the conditioning lesion effect. Our results indicate that better functional recovery is already apparent one week after nerve implantation, and it is due to the conditioning lesion effect, since tenotomy prevents such improvement. The tenotomy effect underlines the fact that some environmental factors concerning the target tissue, and not only the predegenerated nerve, are involved in the conditioning effect. © 1995 Wiley‐Liss, I

ISSN:0360-4012    

DOI:10.1002/jnr.490400308

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThe nature of the axon signal for the induction of proliferation and differentiation of peripheral glial cells is still unknown. Besides the existence of interactions among surface molecules the cellular responses can also be regulated by physicochemical parameters of the membrane. We have previously reported that planar axolemma monolayers coated on glass coverslips at different defined surface molecular packing affected the Schwann cell (SC) morphology and their proliferative response (Calderon et al.: J Neurosci Res 34:206–218, 1993). In this paper we report that relative to SC cultured on uncoated coverslips, the level of expression of both glycoprotein P0and galactocerebroside (GC) (as revealed by immunofluorescence) was increased 2–4 times in SC cultured on axolemma monolayers with either high or low molecular packing. However, the cellular distribution of these antigens was profoundly influenced by the molecular packing density of the axolemma monolayer. SC cultured on an axolemma monolayer at high molecular packing showed preferential expression of P0at the SC surface whereas GC was concentrated intracellularly. On the other hand, SC grown on an axolemma monolayer at low molecular density GC showed preferential expression at the cell surface whereas P0was concentrated intracellularly. © 1995 Wiley‐Lis

ISSN:0360-4012    

DOI:10.1002/jnr.490400309

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractPrevious studies have shown that the mechanical wounding of 3‐week‐old cultured rat astrocytes results in cell proliferation and hypertrophy resembling astrocyte responses to a brain injury in vivo. We now report the effects of basic fibroblast growth factor (bFGF) and an anti‐bFGF antibody on astrocyte morphology, proliferation, and migration following in vitro wounding of confluent secondary cultures. Addition of bFGF (20 ng/ml) to wounded cultures induced morphological changes characteristic of differentiation in wounded and nonwounded areas of the culture. Combined treatment with bFGF and an anti‐bFGF antibody (100 μg/ml) prevented this effect. Astrocyte proliferation along the edges of a scratch wound was at maximum 24 hr after wounding in cells growing in Eagle's minimum essential medium (EMEM) containing 10% serum. Low serum concentration and treatment with dibutyryl cyclic adenosine monophosphate (dbc‐AMP) reduced injury‐associated astrocyte proliferation. Addition of bFGF to cultures in EMEM with serum increased astrocyte proliferation at 18 and 24 hr after wounding. This effect was reduced considerably by treatment of cultures with bFGF in combination with an anti‐bFGF antibody. The combined treatment and the antibody alone reduced cell division to a level lower than in control cultures. Twenty‐four hr following wounding, astrocytes along the edges of the wound exhibited extension of thick, flat processes into the wound area. At 3 and 5 days after wounding, a bodily migration of astrocytes into the wounded area was observed. Addition of bFGF significantly increased astrocyte migration 1 day after wounding, with maximum effect on day 3 and no subsequent increase on day 5. A combination of bFGF and anti‐bFGF antibody as well as the antibody alone reduced astrocyte migration to a level lower than in controls. Immunohistochemical localization and isoform pattern of bFGF in astrocytes did not change with dbc‐AMP treatment or wounding. We conclude that mechanically wounded confluent astrocytes respond to bFGF added to the culture medium by enhancing cell division, differentiation, and migration. In addition, the results of the antibody treatment also suggest a role for endogenous bFGF in astrocyte proliferation and migration elicited by wounding in vitro. These results support the notion that in vivo, both bFGF released by injury and endogenous bFGF synthesized by astrocytes, contribute to the cellular responses that lead to astrogliosis. ©

ISSN:0360-4012    

DOI:10.1002/jnr.490400310

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractWe used the patch clamp technique to study the effect of acetyl‐L‐carnitine arginyl amide (ALCAA) and of nerve growth factor (NGF) on availability of L‐type Ca2+channels in rat pheochromocytoma (PC12) cells maintained in defined medium. Channel availability was measured as number of channels in the patch × the probability of opening (n.P0). In patches from control cells, cells exposed to NGF (10 ng/ml) for six days, and cells exposed to ALCAA (1 mM) for six days, n.P0, measured during 200–240 ms pulses to ‐10 mV (holding potential, −60 mV), was 0.102 ± 0.089 (5 cells), 0.173 ± 0.083 (5 cells), and 0.443 ± 0.261 (7 cells), respectively. The 4.3‐fold increase for the ALCAA‐treated cells was significantly different from control (P<0.05), whereas that for the NGF‐treated cells was not. For the same conditions, the maximum number of superimposed openings at −10 mV was 1.3 ± 0.5 (6 cells), 1.6 ± 0.5 (8 cells), and 3.3 ± 1.8 (8 cells), with the value for the ALCAA‐treated cells being significantly different from control (P<0.001). Additional analysis showed that the distribution of channel open times, the time constants, and the voltage dependence of activation were not changed by prolonged exposure to ALCAA. Short‐term exposure to both ALCAA as well as to the parent compound, acetyl‐L‐carnitine (ALCAR), did not cause an increase but rather a decrease in n.P0, and this short‐term effect of both compounds was blocked by neomycin, an inhibitor of phospholipase C. Together, our findings are consistent with the interpretation that short‐term exposure to ALCAA inhibits Ca2+channel activity, possibly by increasing intracellular Ca2+, and that long‐term exposure causes an increase in Ca2+channel density, possibly by increasing channel expression, with no change in Ca2+ch

ISSN:0360-4012    

DOI:10.1002/jnr.490400311

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY