Expression and androgen regulation of the ciliary neurotrophic factor receptor (CNTFRα) in muscles and spinal cord

We have previously observed that ciliary neurotrophic factor (CNTF) can prevent the degeneration of androgen-sensitive perineal motoneurons and their target muscles, the bulbocavernosus and levator ani (BC/LA), in perinatal female rats. Response to CNTF is dependent on the expression of the alpha component of the CNTF receptor (CNTFRα). In the present study, we examined the developmental profile and androgen regulation of CNTFRα gene expression in BC/LA muscle, thigh muscle, and lumbosacral spinal cord. CNTFRα mRNA was abundantly expressed in the BC/LA and thigh around the time of birth; expression declined progressively after birth and remained low into adulthood. In contrast, CNTFRα message remained high in the lumbosacral spinal cord throughout development. Androgen regulation of CNTFRα expression was examined in prenatal animals by administering the androgen receptor blocker hydroxyflutamide from embryonic days E18 through E21. Four days of androgen deprivation caused a significant up-regulation of CNTFRα mRNA in the BC/LA, thigh, and spinal cord of male fetuses. After castration in adulthood, CNTFRα expression in the BC/LA transiently increased, then decreased below control levels. Expression of CNTFRα in thigh muscles and the lumbosacral spinal cord was not affected by adult castration. Thus, the perineal muscles and motoneurons are potential sites of direct CNTF action, and expression of the CNTFRα gene is modulated by androgen, especially in the androgen-sensitive perineal muscles. Transient up-regulation of CNTFRα following castration or androgen receptor blockade may represent a protective response designed to counteract the muscle atrophy normally induced by androgen withdrawal. © 1998 John Wiley & Sons, Inc. J Neurobiol 35: 217–225, 1998     

Expression of CNTF/LIF‐receptor components and activation of STAT3 signaling in axotomized facial motoneurons: Evidence for a sequential postlesional function of the cytokines

Several lines of evidence suggest that ciliary neurotrophic factor (CNTF) and leukemia inhibitory factor (LIF) are important for the survival and regeneration of axotomized motoneurons. To investigate the role of CNTF/LIF signaling in regenerative responses of motoneurons, we studied the expression of the three receptor components, CNTF receptor α (CNTFRα), LIF receptor β (LIFRβ), and gp130, and the activation of the STAT3 signal transduction pathway in the rat facial nucleus following peripheral nerve transection. As shown by in situ hybridization and immunoblotting, axotomy resulted in a rapid down‐regulation of CNTFRα mRNA expression within 24 h and a concomitant massive up‐regulation of LIFRβ mRNA and protein in the lesioned motoneurons. The altered mRNA levels were maintained for 3 weeks but had returned back to control levels by 6 weeks postlesion after successful regeneration. In contrast, mRNA levels remained in the lesioned state during the 6‐week period studied, when regeneration was prevented by nerve resection. Significant lesion‐induced changes in gp130 mRNA levels were not detectable. Rapid (within 24 h) and sustained (for at least 5 days) activation of STAT3 in axotomized facial motoneurons was revealed by demonstrating the phosphorylation and nuclear translocation of the protein using immunocytochemistry and immunoblotting. In agreement with previous studies showing a complementary regulation of CNTF and LIF in the lesioned facial nerve, our observations on the postlesional regulation of CNTF/LIF receptor components in the facial nucleus indicate a direct and sequential action of the two neurotrophic proteins on axotomized facial motoneurons. © 1999 John Wiley & Sons, Inc. J Neurobiol 41: 559–571, 1999     

Regulation of clusterin expression following spinal cord injury

We have investigated the localization and regulation of a putative extracellular chaperone, clusterin, in the rat spinal cord after lesion. In control animals, clusterin is expressed in motoneurons, in meningeal and ependymal cells, and in astrocytes mainly located beneath the pial surface. Beginning at day 2 after hemisection at segmental level C6, clusterin levels increase in GFAP-positive astrocytes within the lesioned segment. Three weeks after trauma, clusterin mRNA and protein are elevated in neurons close to the lesion site and in glial elements within scar tissue and within degenerating fiber tracts rostral and caudal to the lesion. This study provides evidence for a role of clusterin in the subacute and late phase of spinal cord injury.     

Expression and Cellular Distribution of Ubiquitin in Response to Injury in the Developing Spinal Cord of Monodelphis domestica

Ubiquitin, an 8.5 kDa protein associated with the proteasome degradation pathway has been recently identified as differentially expressed in segment of cord caudal to site of injury in developing spinal cord. Here we describe ubiquitin expression and cellular distribution in spinal cord up to postnatal day P35 in control opossums (Monodelphis domestica) and in response to complete spinal transection (T10) at P7, when axonal growth through site of injury occurs, and P28 when this is no longer possible. Cords were collected 1 or 7 days after injury, with age-matched controls and segments rostral to lesion were studied. Following spinal injury ubiquitin levels (western blotting) appeared reduced compared to controls especially one day after injury at P28. In contrast, after injury mRNA expression (qRT-PCR) was slightly increased at P7 but decreased at P28. Changes in isoelectric point of separated ubiquitin indicated possible post-translational modifications. Cellular distribution demonstrated a developmental shift between earliest (P8) and latest (P35) ages examined, from a predominantly cytoplasmic immunoreactivity to a nuclear expression; staining level and shift to nuclear staining was more pronounced following injury, except 7 days after transection at P28. After injury at P7 immunostaining increased in neurons and additionally in oligodendrocytes at P28. Mass spectrometry showed two ubiquitin bands; the heavier was identified as a fusion product, likely to be an ubiquitin precursor. Apparent changes in ubiquitin expression and cellular distribution in development and response to spinal injury suggest an intricate regulatory system that modulates these responses which, when better understood, may lead to potential therapeutic targets.     

A transient role for ciliary neurotrophic factor in chick photoreceptor development

Previous studies suggest that ciliary neurotrophic factor (CNTF) may represent one of the extrinsic signals controlling the development of vertebrate retinal photoreceptors. In dissociated cultures from embryonic chick retina, exogenously applied CNTF has been shown to act on postmitotic rod precursor cells, resulting in an two- to fourfold increase in the number of cells acquiring an opsin-positive phenotype. We now demonstrate that the responsiveness of photoreceptor precursors to CNTF is confined to a brief phase between their final mitosis and their terminal differentiation owing to the temporally restricted expression of the CNTF receptor (CNTFRα). As shown immunocytochemically, CNTFRα expression in the presumptive photoreceptor layer of the chick retina starts at embryonic day 8 (E8) and is rapidly down-regulated a few days later prior to the differentiation of opsin-positive photoreceptors, both in vivo and in dissociated cultures from E8. We further show that the CNTF-dependent in vitro differentiation of rods is followed by a phase of photoreceptor-specific apoptotic cell death. The loss of differentiated rods during this apoptotic phase can be prevented by micromolar concentrations of retinol. Our results provide evidence that photoreceptor development depends on the sequential action of different extrinsic signals. The time course of CNTFRα expression and the in vitro effects suggest that CNTF or a related molecule is required during early stages of rod differentiation, while differentiated rods depend on additional protective factors for survival. © 1998 John Wiley & Sons, Inc. J Neurobiol 37: 672–683, 1998     

Expression of nerve growth factor receptor mRNA is developmentally regulated and increased after axotomy in rat spinal cord motoneurons

In situ hybridization histochemistry and RNA blot analysis were used to study expression of nerve growth factor receptor (NGF-R) mRNA in rat spinal cord motoneurons. The results show that NGF-R mRNA is expressed at high levels in rat spinal cord motoneurons at the time of naturally occurring cell death. This expression is sustained, but reduced, during synapse formation and is subsequently greatly reduced in the adult spinal cord. A unilateral crush lesion of the sciatic nerve resulted in an 8-fold increase in NGF-R mRNA in adult rat spinal cord motoneurons 3 days after lesion, compared with the nonlesioned side. NGF-R mRNA induction was even more pronounced 7 and 14 days after lesion, reaching levels 12 times higher than those on the nonlesioned side. However, 6 weeks after lesion, when the motor function of the leg was largely restored, NGF-R expression had decreased to levels similar to those on the contralateral side. We therefore suggest that NGF-R mediates a trophic or axonal guidance function for developing and regenerating spinal cord motoneurons.     

Effects of testosterone on the development of a sexually dimorphic neuromuscular system in ciliary neurotrophic factor receptor knockout mice

Motoneurons in the spinal nucleus of the bulbocavernosus (SNB) innervate the perineal muscles, bulbocavernosus (BC), and levator ani (LA). Testosterone regulates the survival of SNB motoneurons and BC/LA muscles during perinatal life. Previous findings suggest that effects of testosterone on this system may be mediated by trophic factors—in particular, by a factor acting through the ciliary neurotrophic factor α‐receptor (CNTFRα). To test the role of CNTFRα in the response of the developing SNB system to testosterone, CNTFRα +/+ and −/− mice were treated with testosterone propionate (TP) or oil during late embryonic development. BC/LA muscle size and SNB motoneuron number were evaluated on the day of birth. Large sex differences in BC and LA muscle size were present in newborn mice of both genotypes, but muscle volumes were reduced in CNTFRα −/− animals relative to same‐sex, wild‐type controls. Prenatal testosterone treatment completely eliminated the sex difference in BC/LA muscle size in wild‐type animals, and eliminated the effect of the CNTFRα gene deletion on muscle size in males. However, the effect of TP treatment on BC and LA muscle sizes was blunted in CNTFRα −/− females. SNB motoneuron number was sexually dimorphic in oil‐treated, wild‐type mice. In contrast, there was no sex difference in SNB motoneuron number in oil‐treated, CNTFRα knockout mice. Prenatal treatment with testosterone did not increase SNB motoneuron number in CNTFRα −/− mice, but also did not significantly increase SNB motoneuron number in newborn wild‐type animals. These findings confirm the absence of a sex difference in SNB motoneuron number in CNTFRα −/− mice. Moreover, the CNTFRα gene deletion influences perineal muscle development and the response of the perineal muscles to testosterone. Prenatal TP treatment of CNTFRα −/− males overcomes the effects of the gene deletion on the BC and LA muscles without a concomitant effect on SNB motoneuron number. © 1999 John Wiley & Sons, Inc. J Neurobiol 41: 317–325, 1999     

Early administration of methylprednisolone decreases apoptotic cell death after spinal cord injury

The purpose of this study is to evaluate, in an experimental model of spinal cord injury (SCI), the presence of apoptotic cell death after trauma and if early administration of a single bolus of methylprednisolone (MP) influences apoptosis in the zone of trauma and in adjacent spinal cord segments. For this study, a total of 96 adult female Wistar rats were subjected to spinal contusion at the T6-T8 level, producing immediate paraplegia. Forty-eight animals (treated group) received a single intraperitoneal injection of MP, at a dose of 30 mg/kg body weight, 10 minutes later. Cells undergoing apoptosis were detected by means of immunohistochemical labeling with the monoclonal antibody Apostain (anti-ssDNA MAb F7-26), in the injured spinal cord tissue, both in the zone of the lesion and in the adjacent spinal segments (rostral and caudal zones), 1, 4, 8, 24 and 72 hours and 1 week after injury. Apoptosis was detected in neurons and glial cells in the zone of the lesion 1 hour after trauma, with a pattern that showed no changes 4 hours later. Between 4 and 8 hours postinjury, the number of apoptotic cells increased, after which it decreased over the following days. In the adjacent spinal segments, apoptotic cells were detected 4 hours after trauma, and increased progressively over the remainder of the study, the number of apoptotic cells being similar in the lesion zone and in rostral and caudal zones one week after injury. When the group of MP-treated animals was considered, significant decreases in the number of apoptotic cells were detected in the lesion zone 24 hours after injury, and in the rostral and caudal zones, at 72 hours and at 1 week after trauma. These findings show that early administration of a single bolus of MP decreases apoptotic cell death after SCI, supporting the utility of MP in reducing secondary damage in injured spinal cord tissue.     

Alterations in the expression of the apurinic/apyrimidinic endonuclease-1/redox factor-1 (APE/ref-1) and DNA damage in the caudal region of acute and chronic spinal cord injured rats treated by embryonic neural stem cells

The oxidative mechanisms of injury-induced damage of neurons within the spinal cord are not very well understood. We used a model of T8-T9 spinal cord injury (SCI) in the rat to induce neuronal degeneration. In this spinal cord injury model, unilateral avulsion of the spinal cord causes oxidative stress of neurons. We tested the hypothesis that apurinic/apyrimidinic endonuclease (or redox effector factor-1, APE/Ref-1) regulates this neuronal oxidation mechanism in the spinal cord region caudal to the lesion, and that DNA damage is an early upstream signal. The embryonic neural stem cell therapy significantly decreased DNA-damage levels in both study groups - acutely (followed up to 7 days after SCI), and chronically (followed up to 28 days after SCI) injured animals. Meanwhile, mRNA levels of APE/Ref-1 significantly increased after embryonic neural stem cell therapy in acutely and chronically injured animals when compared to acute and chronic sham groups. Our data has demonstrated that an increase of APE/Ref-1 mRNA levels in the caudal region of spinal cord strongly correlated with DNA damage after traumatic spinal cord injury. We suggest that DNA damage can be observed both in lesional and caudal regions of the acutely and chronically injured groups, but DNA damage is reduced with embryonic neural stem cell therapy.     

Structural and functional regeneration after spinal cord injury in the weakly electric teleost fish, <Emphasis Type="Italic">Apteronotus</Emphasis><Emphasis Type="Italic">leptorhynchus</Emphasis>

In contrast to mammals, teleost fish exhibit an enormous potential to regenerate adult spinal cord tissue after injury. However, the mechanisms mediating this ability are largely unknown. Here, we analyzed the major processes underlying structural and functional regeneration after amputation of the caudal portion of the spinal cord in Apteronotus leptorhynchus, a weakly electric teleost. After a transient wave of apoptotic cell death, cell proliferation started to increase 5 days after the lesion and persisted at high levels for at least 50 days. New cells differentiated into neurons, glia, and ependymal cells. Retrograde tract tracing revealed axonal re-growth and innervation of the regenerate. Functional regeneration was demonstrated by recovery of the amplitude of the electric organ discharge, a behavior generated by spinal motoneurons. Computer simulations indicated that the observed rates of apoptotic cell death and cell proliferation can adequately explain the re-growth of the spinal cord. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Tail regeneration and ependymal outgrowth in the adult newt, Notophthalmus viridescens, are adversely affected by experimentally produced ischemia

Spinal axons of the adult newt will regenerate when the spinal cord is severed or when the tail is amputated. Ischemia and associated hypoxia have been correlated with poor central nervous system regeneration in mammals. To test the effects of ischemia on newt spinal cord regeneration, the spinal cord and major blood vessels of the newt tail were severed 2 cm caudal to the cloaca as a primary injury. This primary injury severely reduced circulation in the caudal direction for 7 days; by day 8, circulation was largely restored. After various periods of time after primary injury, tails were amputated 1 cm caudal to the primary injury (in the area of ischemia) and tested for regeneration. If the tail was amputated within 5 days of the primary injury, regeneration did not occur. If amputation was 7 days or longer after the primary injury, a regenerative response occurred. Histology showed that in the non-regenerating tails the spinal cord and associated ependyma, known to be important to tail regeneration, had degenerated in the rostral direction. Such degeneration was prevented when tails were first amputated and allowed to form blastemas before the primary injury. The data indicate that the first 5-7 days of blastema formation are particularly sensitive to compromised blood flow (ischemia/hypoxia). It follows that mechanisms must be present in the adult newt to reduce ischemia to a minimum and thus allow ependymal outgrowth and tail regeneration.     

Upregulation of vascular endothelial growth factor receptors Flt-1 and Flk-1 following acute spinal cord contusion in rats.

To investigate the possible role of vascular endothelial growth factor (VEGF) in the injured spinal cord, we analyzed the distribution and time course of the two tyrosine kinase receptors for VEGF, Flt-1 and Flk-1, in the rat spinal cord following contusion injury using a weight-drop impactor. The semi-quantitative RT-PCR analysis of Flt-1 and Flk-1 in the spinal cord showed slight upregulation of these receptors following spinal cord injury. Although mRNAs for Flt-1 and Flk-1 were constitutively expressed in neurons, vascular endothelial cells, and some astrocytes in laminectomy control rats, their upregulation was induced in association with microglia/macrophages and reactive astrocytes in the vicinity of the lesion within 1 day in rats with a contusion injury and persisted for at least 14 days. The spatiotemporal expression of Flt-1 in the contused spinal cord mirrored that of Flk-1 expression. In the early phase of spinal cord injury, upregulation of Flt-1 and Flk-1 mRNA occurred in microglia/macrophages that infiltrated the lesion. In addition, the expression of both receptors increased progressively in reactive astrocytes within the vicinity of the lesion, predominately in the white matter, and almost all reactive astrocytes coexpressed Flt-1 or Flk-1 and nestin. These results suggest that VEGF may be involved in the inflammatory response and the astroglial reaction to contusion injuries of the spinal cord via specific VEGF receptors.     

Succinate dehydrogenase activity in rat dorsolateral ventral horn motoneurons at L6 after spaceflight and recovery

Succinate dehydrogenase (SDH) activity levels of motoneurons in the rostral, middle, and caudal portions of the dorsolateral region of the ventral horn of the 6th lumbar (L6) segment of the rat spinal cord were determined after 14 days of spaceflight and after 9 days of recovery on Earth. The mean SDH activity of motoneurons with cell body sizes between 500 and 800 micrometers2 located in the rostral portion of the L6 segment was lower in spaceflight than in age-matched control rats. The decrease in motoneuron SDH activity persisted for at least 9 days of recovery on Earth. In contrast, the mean SDH activity of motoneurons located in the middle and caudal portions of the L6 segment were unaffected by spaceflight and recovery on Earth. The motoneurons in the rostral portion of the L6 segment presumably innervate both high- and low-oxidative fibers in hindlimb muscles, whereas those in the middle and caudal portions presumably innervate perineal muscles that are comprised only of low-oxidative fibers. These data indicate that moderate-sized motoneurons, most likely innervating fibers in high-oxidative muscles, are responsive to the microgravity environment.     

Changes in NMDA receptor subunit expression in response to contusive spinal cord injury

Differential assembly of N-methyl-D-aspartate (NMDA) receptor subunits determines their functional characteristics. Using in situ hybridization, we found a selective increase of the subunits NR1 and NR2A mRNA at 24 h in ventral motor neurons (VMN) caudal to a standardized spinal cord contusion injury (SCI). Other neuronal cell populations and VMN rostral to the injury site appeared unaffected. Significant up-regulation of NR2A mRNA also was seen 1 month after SCI in thoracic and lumbar VMN. The selective effects on VMN caudal to the injury site suggest that the loss of descending innervation leads to increased NMDA receptor subunit expression in these cells after SCI, which may alter their responses to glutamate. In contrast, protein levels determined by western blot analysis show decreased levels of NR2A 1 month after SCI in whole thoracic segments of spinal cord that included the injury sites. No effects of injury were seen on subunit levels in cervical or lumbar segments. Taken together with our previous study showing alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor subunit down-regulation after injury, our data suggest that glutamate receptor composition is significantly altered after SCI. These changes need to be taken into account to properly understand the function of, and potential pharmacotherapy for, the chronically injured spinal cord.     

Aberrant Crossed Corticospinal Facilitation in Muscles Distant from a Spinal Cord Injury

Crossed facilitatory interactions in the corticospinal pathway are impaired in humans with chronic incomplete spinal cord injury (SCI). The extent to which crossed facilitation is affected in muscles above and below the injury remains unknown. To address this question we tested 51 patients with neurological injuries between C2-T12 and 17 age-matched healthy controls. Using transcranial magnetic stimulation we elicited motor evoked potentials (MEPs) in the resting first dorsal interosseous, biceps brachii, and tibialis anterior muscles when the contralateral side remained at rest or performed 70% of maximal voluntary contraction (MVC) into index finger abduction, elbow flexion, and ankle dorsiflexion, respectively. By testing MEPs in muscles with motoneurons located at different spinal cord segments we were able to relate the neurological level of injury to be above, at, or below the location of the motoneurons of the muscle tested. We demonstrate that in patients the size of MEPs was increased to a similar extent as in controls in muscles above the injury during 70% of MVC compared to rest. MEPs remained unchanged in muscles at and within 5 segments below the injury during 70% of MVC compared to rest. However, in muscles beyond 5 segments below the injury the size of MEPs increased similar to controls and was aberrantly high, 2-fold above controls, in muscles distant (>15 segments) from the injury. These aberrantly large MEPs were accompanied by larger F-wave amplitudes compared to controls. Thus, our findings support the view that corticospinal degeneration does not spread rostral to the lesion, and highlights the potential of caudal regions distant from an injury to facilitate residual corticospinal output after SCI.     

Spatiotemporal Patterns of SSeCKS Expression After Rat Spinal Cord Injury

Src suppressed C kinase substrate (SSeCKS) was identified as a PKC substrate/PKC-binding protein, which plays a role in mitogenic regulatory activity and has a function in the control of cell signaling and cytoskeletal arrangement. However its distribution and function in the central nervous system (CNS) lesion remain unclear. In this study, we mainly investigated the mRNA and protein expression and cellular localization of SSeCKS during spinal cord injury (SCI). Real-time PCR and Western blot analysis revealed that SSeCKS was present in normal whole spinal cord. It gradually increased, reached a peak at 3 days for its mRNA level and 5 days for its protein level after SCI, and then declined during the following days. In ventral horn, the expression of SSeCKS underwent a temporal pattern that was similar with the whole spinal cord in both mRNA and protein level. However, in dorsal horn, the mRNA and protein for SSeCKS expression were significantly increased at 1 day for its mRNA level and 3 days for its protein level, and then gradually declined to the baseline level, ultimately up-regulated again from 7 to 14 days. The protein expression of SSeCKS was further analysed by immunohistochemistry. The positively stained areas for SSeCKS changed with the similar pattern to that of protein expression detected by immunoblotting analysis. Double immunofluorescence staining showed that SSeCKS immunoreactivity (IR) was found in neurons, astrocytes, oligodendrocytes of spinal cord tissues within 5 mm from the lesion site. Importantly, injury-induced expression of SSeCKS was co-labeled by active caspase-3 (apoptotic marker), Tau-1 (the marker for pathological oligodendrocyte) and β-1,4-galactosyltransferase 1 (GalT). All the results suggested that SSeCKS might play important roles in spinal cord pathophysiology and further research is needed to have a good understanding of its function and mechanism. Feng Xiao and Min Fei contributed equally to this work.     

Anatomical and functional recovery folloing spinal cord transection in the chick embryo

Following complete transection of the thoracic spinal cord at various times during embryonic development, chick embryos and posthatched animals exhibited various degrees of anatomical and functional recovery depending upon the age of injury. Transection on embryonic day 2 (E2), when neurogenesis is still occurring and before descending or ascending fiber tracts have formed, produced no noticeable behavioral or anatomical deficits. Embryos hatched on their own and were behaviorally indistinguishable from control hatchlings. Similar results were found following transection on E5, an age when neurogenesis is complete and when ascending and descending fiber tracts have begun to project through the thoracic region. Within 48 h following injury on E5, large numbers of nerve fibers were observed growing across the site of transection. By E8, injections of horseradish peroxidase (HRP) administered caudal to the lesion, retrogradely labelled rostral spinal and brainstem neurons. Embryos transected on E5 were able to hatch and could stand and locomote posthatching in a manner that was indistinguishable from controls. Following spinal cord transections on E10, anatomical recovery of the spinal cord at the site of injury was not quite as complete as after E5 transection. Nonetheless, anatomical continuity was restored at the site of injury, axons projected across this region, and rostral spinal and brainstem neurons could be retogradely labelled following HRP injections administered caudal to the lesion. At least part of this anatomical recovery may be mediated by the regeneration or regrowth of lesioned axons. Although none of the embryos transected on E10 that survived to hatching were able to hatch on their own, because several shamoperated embryos were also unable to hatch, we do not attribute this deficit to the spinal transection. When E10-transected embryos were aided in escaping from the shell, they were able to support their own weight, could stand, and locomote, and were generally comparable, behaviorally, to control hatchlings. Repair of the spinal cord following transection on E15 was considerably less complete compared to embryos transected on E2, E5, or E10. However, in some cases, a degree of anatomical continuity was eventually restored and a few spinal neurons rostral to the lesion could be retrogradely labelled with HRP. By contrast, labelled brainstem neurons were never observed following E15 transection. E15 transected embryos were never able to hatch on their own, and when aided in escaping from the shell, the hatchlings were never able to stand, support their own weight or locomote. We conclude that successful anatomical and functional recovery occurs following a complete spinal cord transection in the chick embryo made any time between E2 and E10. By E15, however, there is an altered response to the transection such that anatomical continuity is not restored sufficiently to mediate behavioral or functional recovery. Although the altered response of the chick embryo spinal cord to injury between E10 and E15 could be due to a variety of factors, we favor the notion that cellular or molecular changes associated with axonal growth and guidance occur at this time that are responsible for the transition from successful to unsuccessful recovery.     

Ciliary neurotrophic factor protects retinal ganglion cells from axotomy‐induced apoptosis via modulation of retinal glia in vivo

Adenoviral‐mediated transfer of ciliary neurotrophic factor (CNTF) to the retina rescued retinal ganglion cells (RGCs) from axotomy‐induced apoptosis, presumably via activation of the high affinity CNTF receptor alpha (CNTFRα) expressed on RGCs. CNTF can also activate astrocytes, via its low affinity leukemia inhibitory receptor beta expressed on mature astrocytes, suggesting that CNTF may also protect injured neurons indirectly by modulating glia. Adenoviral‐mediated overexpression of CNTF in normal and axotomized rat retinas was examined to determine if it could increase the expression of several glial markers previously demonstrated to have a neuroprotective function in the injured brain and retina. Using Western blotting, the expression of glial fibrillary acid protein (GFAP), glutamate/aspartate transporter‐1 (GLAST‐1), glutamine synthetase (GS), and connexin 43 (Cx43) was examined 7 days after intravitreal injections of Ad.CNTF or control Ad.LacZ. Compared to controls, intravitreal injection of Ad.CNTF led to significant changes in the expression of CNTFRα, pSTAT₃, GFAP, GLAST, GS, and Cx43 in normal and axotomized retinas. Taken together, these results suggest that the neuroprotective effects of CNTF may result from a shift of retinal glia cells to a more neuroprotective phenotype. Moreover, the modulation of astrocytes may buffer high concentrations of glutamate that have been shown to contribute to the death of RGCs after optic nerve transection. © 2005 Wiley Periodicals, Inc. J Neurobiol, 2005     

Neurotrophic agents prevent motoneuron death following sciatic nerve section in the neonatal mouse

We have examined the ability of different neurotrophic and growth factors to prevent axotomy-induced motoneuron cell death in the developing mouse spinal cord. After postnatal unilateral section of the mouse sciatic nerve, most motoneuron (MN) loss occurs in the lateral motor column of the fourth lumbar segment (L4). Significant axotomy-induced cell death occurred after surgery performed on or before postnatal day (PN) 5. In contrast, no significant cell loss was found when axotomy was performed after PN10. Axotomy on PN2 or PN5 resulted in a 44% loss of L4 motoneurons by 7 days, and a 66% loss of motoneurons by 10 days postsurgery. Implantation of gelfoam presoaked in various neurotrophic factors at the lesion site rescued axotomized motoneurons. Nerve growth factor (NGF), nedurotrophin-4/5 (NT-4/5) and ciliary neurotrophic factor (CNTF) rescued 20%–30% of motoneurons, whereas brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and insulin-like growth factor 1 (IGF-1) rescued virtually all motoneurons from axotomy-induced death. By contrast, platelet-derived growth factor (PDGF)-AA, PDGF-AB, basic fibroblast growth factor (bFGF), and interleukin (IL-6) were ineffective on motoneuron survival following axotomy. NGF, BDNF, NT-3, IGF-1, and CNTF also prevented axotomy-induced atrophy of surviving motoneurons. These data show that mouse lumbar motoneurons continue to be vulnerable to axotomy up to about 1 week after birth and that a number of trophic agents, including the neurotrophins, CNTF, and IGF-1, can prevent the death of these neurons following axotomy. Our studies confirm and extend previous reports on the time course of axotomy-induced mouse motoneuron death and the survival promoting effects of neurotrophic factors. 1994 John Wiley & Sons, Inc.