Galectin-1 regulates initial axonal growth in peripheral nerves after axotomy. (9/389)

The signals that prompt the axons to send out processes in peripheral nerves after axotomy are not well understood. Here, we report that galectin-1 can play an important role in this initial stage. We developed an in vitro nerve regeneration model that allows us to monitor the initial axon and support cell outgrowth from the proximal nerve stump, which is comparable to the initial stages of nerve repair. We isolated a factor secreted from COS1 cells that enhanced axonal regeneration, and we identified the factor as galectin-1. Recombinant human galectin-1 (rhGAL-1) showed the same activity at low concentrations (50 pg/ml) that are two orders of magnitude lower than those of lectin activity. A similarly low concentration was also effective in in vivo experiments of axonal regeneration with migrating reactive Schwann cells to a grafted silicone tube after transection of adult rat peripheral nerve. Moreover, the application of functional anti-rhGAL-1 antibody strongly inhibited the regeneration in vivo as well as in vitro. The same effect of rhGAL-1 was confirmed in crush/freeze experiments of the adult mouse sciatic nerve. Because galectin-1 is expressed in the regenerating sciatic nerves as well as in both sensory neurons and motor neurons, we suggest that galectin-1 may regulate initial repair after axotomy. This high activity of the factor applied under nonreducing conditions suggests that galectin-1 may work as a cytokine, not as a lectin.  (+info)

Increased spontaneous unit activity and appearance of spontaneous negative potentials in the goldfish tectum during refinement of the optic projection. (10/389)

Spontaneous (not retinally driven) postsynaptic activity was examined during activity-dependent refinement of optic fibers in the goldfish tectum. Unit recordings in vivo and in vitro demonstrated that spontaneous tectal activity increased to 150% of normal during refinement at 1-2 months after optic nerve crush and subsequently returned to baseline over the next month. This increase was not mimicked by long-term denervation indicating an effect specifically influenced by regenerating fibers. Loss of optic input was also found to induce spontaneous negative potentials (SNPs) rapidly in the tectum. SNPs were negative, monophasic potentials of 70-120 msec duration and -0.15 to -1.5 mV amplitude. SNPs occurred with no apparent periodicity at a frequency of approximately 0.3-0.6 Hz. Multiple electrode recordings and depth analysis showed that SNPs were localized events occurring in columnar domains of tectum a few hundred micrometers wide. Cross-correlation analysis revealed that SNPs were strongly correlated with local unit bursting, suggesting SNPs are generated by the summed synaptic and spike currents of coactive cells in small regions of the tectum. SNPs were suppressed by a low concentration of APV indicating they were regulated by NMDA receptors. During regeneration, the number and size of SNPs reached a peak during refinement and subsequently decreased, eventually disappearing. This temporal association with refinement suggests that these patterns of postsynaptic activity may have functional relevance. It is hypothesized that SNPs or the underlying activity that produces them increases the excitability of target cells, allowing the weak, less-convergent input from regenerating axons to drive target groups of cells in the tectum during refinement.  (+info)

Function of skeletal muscle tissue formed after myoblast transplantation into irradiated mouse muscles. (11/389)

1. Pretreatment of muscles with ionising radiation enhances tissue formation by transplanted myoblasts but little is known about the effects on muscle function. We implanted myoblasts from an expanded, male-donor-derived, culture (i28) into X-ray irradiated (16 Gy) or irradiated and damaged soleus muscles of female syngeneic mice (Balb/c). Three to 6 months later the isometric contractile properties of the muscles were studied in vitro, and donor nuclei were visualised in muscle sections with a Y chromosome-specific DNA probe. 2. Irradiated sham-injected muscles had smaller masses than untreated solei and produced less twitch and tetanic force (all by about 18 %). Injection of 106 myoblasts abolished these deficiencies and innervation appeared normal. 3. Cryodamage of irradiated solei produced muscle remnants with few (1-50) or no fibres. Additional myoblast implantation led to formation of large muscles (25 % above normal) containing numerous small-diameter fibres. Upon direct electrical stimulation, these muscles produced considerable twitch (53 % of normal) and tetanic forces (35 % of normal) but innervation was insufficient as indicated by weak nerve-evoked contractions and elevated ACh sensitivity. 4. In control experiments on irradiated muscles, reinnervation was found to be less complete after botulinum toxin paralysis than after nerve crush indicating that proliferative arrest of irradiated Schwann cells may account for the observed innervation deficits. 5. Irradiation appears to be an effective pretreatment for improving myoblast transplantation. The injected cells can even produce organised contractile tissue replacing whole muscle. However, impaired nerve regeneration limits the functional performance of the new muscle.  (+info)

Nitric oxide influences injury-induced microglial migration and accumulation in the leech CNS. (12/389)

Damage to the leech or mammalian CNS increases nitric oxide (NO) production and causes accumulation of phagocytic microglial cells at the injury site. The aim of this study was to determine whether NO plays a role in microglial migration and accumulation at lesions in which NO is generated by a rapidly appearing endothelial nitric oxide synthase (eNOS) in leeches. Immunohistochemistry and cytochemistry demonstrated active eNOS before and throughout the period of microglial accumulation at the lesion. Decreasing NO synthesis by application of the NOS inhibitor N(w)-nitro-L-arginine methyl ester (1 mM) significantly reduced microglial accumulation, whereas its inactive enantiomer N(w)-nitro-D-arginine methyl ester (1 mM) resulted in microglial accumulation similar to that in crushed controls. Increasing NO with the donor spermine NONOate (SPNO) (1 mM) also inhibited accumulation, but not in the presence of the NO scavenger 2-(4-carboxyphenyl)-4,4,5, 5-teramethylimidazoline-oxyl-3-oxide (50 microM). The effect of SPNO was reversed by washout. SPNO application reduced average microglial migratory speeds and even reversibly arrested cell movement, as measured in living nerve cords. These results suggest that NO produced at a lesion may be a stop signal for microglia to accumulate there and that it can act on microglia early in their migration. Thus, NO may assume a larger role in nerve repair and recovery from injury by modulating accumulation of microglia, which appear to be important for axonal regeneration.  (+info)

The effects of FK506 on retinal ganglion cells after optic nerve crush. (13/389)

PURPOSE: The purpose of this study was twofold: to determine whether immunophilins were present in the rat retina and to determine the physiologic consequence of their presence. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis were performed on rat retinal tissue, and the immunophilin FKBP12 was found to be present in retina. Immunohistochemical studies showed the presence of FKBP12 in retinal ganglion cells (RGCs). In rats, optic nerve crush was performed on one side and a sham operation on the other side. By gavage, animals were given 5 mg/kg per day of the FKBP12 ligand FK506 in sterile phosphate-buffered saline (PBS) or in PBS alone. Eight days after nerve crush, the total number of back-labeled RGCs was estimated from retinal wholemounts. RESULTS: In control eyes, the number of labeled ganglion cells was 74,104 +/- 4,166 (mean +/- SEM) in rats receiving vehicle and 74,993 +/- 3,098 in animals receiving FK506 daily. Eight days after optic nerve crush, 27,775 +/- 3,332 labeled ganglion cells were counted in retinas of animals receiving vehicle (n = 11), whereas 33% more ganglion cells (37,118 +/- 2,475) were counted in animals receiving FK506 daily (n = 11). This difference was statistically significant (P < 0.05). CONCLUSIONS: The data presented demonstrate that the immunophilin FKBP12 is present in retina and specifically in RGCs. In addition, the FKBP12 ligand FK506 confers neuroprotection on RGCs after optic nerve crush. This neuroprotection may occur as a result of FK506's ability to interfere with apoptotic mechanisms after optic nerve crush.  (+info)

Permanent alterations of spinal cord reflexes following nerve lesion in newborn rats. (14/389)

Sciatic nerve lesion in newborn rats is known to cause degeneration of a large number of axotomized motoneurones and spinal ganglion cells. Some of the surviving motoneurones exhibit abnormal firing properties and the projection pattern of central terminals of sensory neurones is altered. We report here on long-term changes in spinal cord reflexes in adult rats following neonatal nerve crush. In acutely spinalized and anaesthetized adult rats 4-6 months old in which the sciatic nerve had been crushed on one side at birth, the tibial nerve, common peroneal nerve or sural nerve were stimulated on the reinnervated and control side and reflex responses were recorded from the L5 ventral spinal roots. Ventral root responses (VRRs) to tibial and peroneal nerve stimulation on the side of the nerve lesion were significantly smaller in amplitude representing only about 15% of the mean amplitude of VRRs on the control side. The calculated central delay of the first, presumably monosynaptic component of the VRR potential was 1.6 ms on the control side while the earliest VRR wave on the side of the nerve lesion appeared after a mean central latency of 4.0 ms that seems too long to be of monosynaptic origin. These results suggest that neonatal sciatic nerve injury markedly alters the physiological properties and synaptic connectivity in spinal cord neurones and causes a marked depression of spinal cord responses to peripheral nerve stimulation.  (+info)

Re-innervation of twitch and slow muscle fibres of the frog after crushing the motor nerves. (15/389)

1. The conduction velocities of individual motor axons innervating twitch and slow muscle fibres of the frog were determined by intracellular recording of junctional potentials elicited by stimulating the motor nerves at two different points. 2. In normal pyriformis muscles twitch and slow fibres were found to be innervated by two distinct populations of motor axons. Twitch fibre axons conducted at 10-18-7 m/sec, while the conduction velocities of slow fibre axons ranged from 0-5 to 5 m/sec (at 7-9 degrees C). The thresholds for electrical stimulation were significantly lower in the fast than in the slow axons population. 3. Following denervation by crushing the sciatic nerve fast axons which re-innervated the muscle had lower conduction velocities than normal but could still be identified. These lower conduction velocities were measured proximal to the site of the crush and did not recover over a period of 446 days. 4. Fast motor axons regenerated more quickly than slow axons and re-innervated twitch as well as slow muscle fibres non-selectively. About 1 month later slow axons re-established synaptic contacts with slow (and some twitch) muscle fibres. Simultaneous re-innervation by fast and slow motor axons was occasionally observed in slow muscle fibres. Finally, the slow muscle fibres were innervated by slow axons only, while synapses of fast axons could no longer be found in this type of muscle fibre. 5. Action potentials were observed in denervated as well as in re-innervated slow muscle fibres; they disappeared as re-innervation progressed. 6. It is concluded that non-selective re-innervation of slow muscle fibres is present in the frog; it is, however, a transient phenomenon followed by restoration of the original innervation pattern.  (+info)

Injury-specific expression of activating transcription factor-3 in retinal ganglion cells and its colocalized expression with phosphorylated c-Jun. (16/389)

PURPOSE: To ascribe activating transcription factor (ATF)-3 as a specifically induced transcription factor after ON injury and to describe its putative role as a modulator of c-Jun transactivation. METHODS: The adult rat optic nerve was crushed intraorbitally, and expression profiles of ATF-3, ATF-2, and phosphorylated c-Jun (p-c-Jun) were examined by immunohistochemistry and ISH. Western blot analysis for ATF-3 and -2 were also performed. Furthermore, colocalized detection of c-Jun mRNA with ATF-2 or -3 was attempted with a combined method of simultaneous immunohistochemistry and in situ hybridization. RESULTS: In response to optic nerve injury, substantial expression of ATF-3 as well as that of p-c-Jun was observed in the retinal ganglion cells, whereas no expression of ATF-3 was seen in other noninjured retinal cells. In contrast, ATF-2 was normally expressed abundantly in both retinal ganglion cells and displaced amacrine cells, but expression dropped in retinal ganglion cells after nerve injury. The expression profiles of ATF-2 and -3 after optic nerve injury were confirmed by Western blot analysis. A higher degree of colocalization was observed for ATF-3 and c-Jun than the modest codetection for ATF-2 and c-Jun. CONCLUSIONS: The transcription factor ATF-3 is specifically induced upon optic nerve injury and colocalizes with p-c-Jun in surviving ganglion cells. These findings suggest that both ATF-3 and c-Jun are crucial to trigger various transcriptional responses and may act synergistically during the survival phase of the optic nerve in the injury model.  (+info)