Heparan sulphate and HB-GAM (heparin-binding growth-associated molecule) in the development of the thalamocortical pathway of rat brain

Extracellular matrix (ECM) molecules, such as laminin, tenascin, chondroitin sulphate proteoglycans and heparan sulphate proteoglycans have been suggested to have 'signpost' and directing roles in the formation of axonal projections in cortical development. We show here that the expression of the neurite outgrowth-promoting protein heparin-binding growth-associated molecule (HB-GAM) and N-syndecan, a transmembrane heparan sulphate proteoglycan previously isolated as a receptor for HB-GAM, is spatiotemporally associated with the developing thalamocortical pathway in the rat brain. Using in situ hybridization, thalamic neurons were shown to express mRNA for N-syndecan, and in vitro, thalamic neurons grew more neurites on HB-GAM than on laminin. The HB-GAM-induced neurite outgrowth in thalamic neurons was inhibited by heparitinase, heparin, soluble N-syndecan and by an excess of soluble HB-GAM in the culture medium. In a pathway assay, thalamic neurons selectively preferred attaching and growing neurites on matrices containing HB-GAM than on those containing fibronectin or laminin alone, suggesting that HB-GAM may modulate the effect of other ECM proteins. On an unfixed brain slice preparation, thalamic neurons repeatedly showed a typical neurite outgrowth and attachment pattern resembling the expression pattern of HB-GAM. On the brain slices, the neurite outgrowth was significantly inhibited by heparitinase, heparin and soluble HB-GAM, thus displaying features of neurite outgrowth on matrix-bound HB-GAM. Our results suggest that HB-GAM is important for the neurite outgrowth of thalamic neurons and it may function as an ECM-bound guidance cue for thalamic neurons that possess HB-GAM-binding heparan sulphates on their cell membrane.     

Preferential Growth of Neonatal Rat Dorsal Root Ganglion Cells on Homotypic Peripheral Nerve Substrates In Vitro

Developing sensory neurons interact with molecular signals in the local environment to generate stereotypic nerve pathways. Regenerating neurons seem to lose the ability to reinnervate their original sites in the targets, resulting in abnormal sensory input and consequent clinical pathophysiology. The specificity of reinnervation of peripheral targets by regenerating axons is thus crucial for normal recovery of function. In this study, we have examined evidence for selectivity of interactions between primary afferent neurons from identified levels of the spinal cord and different peripheral nerve environments by culturing these neurons on sections of nerves to muscle and viscera. We have compared the growth of a population of sensory afferents normally innervating somatic targets (dorsal root ganglion cells from L4 and L5) with populations containing many afferents innervating visceral targets (L6 and S1 dorsal root ganglia and nodose ganglion). These neurons, from newly born rats, were cultured on unfixed cryostat sections of normal and prelesioned gastrocnemius nerve, pelvic spinal nerve and vagus nerve from adult rats. Normal muscle nerve was seen to support the regeneration of a significantly greater proportion of somatic neurons, with longer neurites, than the visceral nerves. Similarly, much higher proportions of the‘visceral’population of afferent neurons were seen to extend neurites on the normal visceral nerve substrates, with longer neurites, than on the muscle nerve substrate. The selectivity displayed by the sensory neurons for their normal nerve substrates was abolished when they were cultured on prelesioned nerve substrates subjected to Wallerian degeneration, which was apparent from the equivalent and increased proportions of growing neurons having comparable neurite lengths, on all the nerve substrates. We conclude that sensory neurons recognize and respond to substrate-specific and substrate-bound molecules present in normal adult peripheral nerves, and that these differences are lost in prelesioned nerves following Wallerian degeneration.     

Developmentally Regulated Expression of HDNF/NT-3 mRNA in Rat Spinal Cord Motoneurons and Expression of BDNF mRNA in Embryonic Dorsal Root Ganglion

Northern blot analysis was used to demonstrate high levels of hippocampus-derived neurotrophic factor/neurotrophin-3 (HDNF/NT-3) mRNA in the embryonic day (E) 13 - 14 and 15 - 16 spinal cord. The level decreased at E18 - 19 and remained the same until postnatal day (P) 1, after which it decreased further to a level below the detection limit in the adult. In situ hybridization revealed that the NT-3 mRNA detected in the developing spinal cord was derived from motoneurons and the decrease seen at E18 - 19 was caused by a reduction in the number of motoneurons expressing NT-3 mRNA. The distribution of NT-3 mRNA-expressing cells in the E15 spinal cord was very similar to the distribution of cells expressing choline acetyltransferase or nerve growth factor receptor (NGFR) mRNA. Moreover, a striking similarity between the developmentally regulated expression of NT-3 and NGFR mRNA was noted in spinal cord motoneurons. A subpopulation of all neurons in the dorsal root ganglia expressed brain-derived neurotrophic factor (BDNF) mRNA from E13, the earliest time examined, to adulthood. These results are consistent with a trophic role of NT-3 for proprioceptive sensory neurons innervating the ventral horn, and imply a local action of BDNF for developing sensory neurons within the dorsal root ganglia.     

Inhibition and enhancement of neural regeneration by chondroitin sulfate proteoglycans

The current dogma in neural regeneration research implies that chondroitin sulfate proteoglycans(CSPGs) inhibit plasticity and regeneration in the adult central nervous system(CNS). We argue that the role of the CSPGs can be reversed from inhibition to activation by developmentally expressed CSPG-binding factors. Heparin-binding growth-associated molecule(HB-GAM; also designated as pleiotrophin) has been studied as a candidate molecule that might modulate the role of CSPG matrices in plasticity and regeneration. Studies in vitro show that in the presence of soluble HB-GAM chondroitin sulfate(CS) chains of CSPGs display an enhancing effect on neurite outgrowth. Based on the in vitro studies, we suggest a model according to which the HB-GAM/CS complex binds to the neuron surface receptor glypican-2, which induces neurite growth. Furthermore, HB-GAM masks the CS binding sites of the neurite outgrowth inhibiting receptor protein tyrosine phosphatase sigma(PTPσ), which may contribute to the HB-GAM-induced regenerative effect. In vivo studies using two-photon imaging after local HB-GAM injection into prick-injury of the cerebral cortex reveal regeneration of dendrites that has not been previously demonstrated after injuries of the mammalian nervous system. In the spinal cord, two-photon imaging displays HB-GAM-induced axonal regeneration. Studies on the HB-GAM/CS mechanism in vitro and in vivo are expected to pave the way for drug development for injuries of brain and spinal cord.     

蛋白激酶C调节脊髓神经元突起的生长(英文)

目的关于蛋白激酶C(PKC)在神经元突起生长和神经再生中的作用,目前仍存有争议。本研究主要观察PKC对离体培养的脊髓神经元生长的调节作用,旨在阐明PKC对突起生长的调节作用。方法分离纯化胎龄14天(E14)的SD胎鼠的脊髓前角神经元,进行原代培养,并检测不同时相点膜/浆PKC活性(m/c-PKCactivity)的比值。结果神经元培养3-11d期间,神经元内m/c-PKC比值以及PKC-βII在突起中的表达水平均与突起生长呈显著相关关系(r=0.95,P<0.01;r=0.73,P<0.01)。此外,PKC激动剂PMA能显著提高m/c-PKC比值,且与神经突起的生长一致(r=0.99,P<0.01)。而PKC抑制剂GF109203X则能显著抑制突起生长,且不被PMA作用所逆转。结论PKC的活性在脊髓神经元突起生长调节中具有重要作用,其中βII亚型可能扮演重要角色。     

Ciliary neurotrophic factor stimulates neurite outgrowth from spinal cord neurons

Ciliary neurotrophic factor (CNTF) has been shown to promote the survival of motoneurons, but its effects on axonal outgrowth have not been examined in detail. Since nerve growth factor (NGF) promotes the outgrowth of neurites within the same populations of neurons that depend on NGF for survival, we investigated whether CNTF would stimulate neurite outgrowth from motoneurons in addition to enhancing their survival. We found that CNTF is a powerful promoter of neurite outgrowth from cultured chick embryo ventral spinal cord neurons. An effect of CNTF on neurite outgrowth was detectable within 7 hours, and at a concentration of 10 ng/ml, CNTF enhanced neurite length by about 3- to 4-fold within 48 hours. The neurite growth-promoting effect of CNTF does not appear to be a consequence of its survival-promoting effect. To determine whether the effect of CNTF on spinal cord neurons was specific for motoneurons, we analyzed cell survival and neurite outgrowth for motoneurons labeled with diI, as well as for neurons taken from the dorsal half of the spinal cord, which lacks motoneurons. We found that the effect of CNTF was about the same for motoneurons as it was for neurons from the dorsal spinal cord. The responsiveness of a variety of spinal cord neurons to CNTF may broaden the appeal of CNTF as a candidate for the treatment of spinal cord injury or disease. © 1996 Wiley-Liss, Inc.     

Raphe-spinal neurons display an age-dependent differential capacity for neurite outgrowth compared to other brainstem-spinal populations

Functional regeneration of brainstem-spinal pathways occurs in the developing chick when the spinal cord is severed prior to embryonic day (E) 13. Functional spinal cord regeneration is not observed in animals injured after E13. This developmental transition from a permissive to a restrictive repair period may be due to the formation of an extrinsic inhibitory environment preventing axonal growth, and/or an intrinsic inability of mature neurons to regenerate. Here, we investigated the capacity of specific populations of brainstem-spinal projection neurons to regrow neurites in vitro from young (E8) versus mature (E17) brainstem explants. A crystal of carbocyanine dye (DiI) was implanted in ovo into the E5 cervical spinal cord to retrogradely label brainstem-spinal projection neurons. Three or 12 days later, discrete regions of the brainstem containing DiI-labeled neurons were dissected to produce explant cultures grown in serum-free media on laminin substrates. The subsequent redistribution of DiI into regenerating processes permitted the study of in vitro neurite outgrowth from identified brainstem-spinal neurons. When explanted on E8, i.e., an age when brainstem-spinal neurons are normally elongating through the spinal cord and are capable of in vivo functional regeneration, robust neurite outgrowth was observed from all brainstem populations, including rubro-, reticulo-, vestibulo-, and raphe-spinal neurons. In contrast, when explanted on E17, robust neurite outgrowth was seen only from raphe-spinal neurons. Neurite outgrowth from raphe-spinal neurons was 5-hydroxy-tryptamine immunoreactive. This study demonstrates that in growth factor-free environments with permissive growth substrates, neurite outgrowth from brainstem-spinal neurons is dependent on both neuronal age and phenotype.     

Translocation Associated Membrane Protein 1 Contributes to Chronic Constriction Injury-Induced Neuropathic Pain in the Dorsal Root Ganglion and Spinal Cord in Rats

Neuropathic pain is a severe debilitating state caused by injury or dysfunction of somatosensory nervous system, and the clinical treatment is still challenging. Translocation associated membrane protein 1 (TRAM1), an adapter protein, participates in a variety of transduction pathways and mediates the biological functions such as cell proliferation, activation, and differentiation. However, whether TRAM1 is involved in the pathogenesis of neuropathic pain is still unclear. In our study, we reported the role of TRAM1 in the maintenance of neuropathic pain induced by chronic constriction injury (CCI) on rats. By western blot and staining, we found that TRAM1 increased in the dorsal root ganglion (DRG) neurons and spinal cord (SC) neurons after CCI. Being similar to IB4-, CGRP-positive expressed area, TRAM1 also expressed in the superficial laminae of the spinal cord dorsal horn (SCDH), suggesting it was related to the innervations of the primary afferents. Moreover, intrathecal injection of TRAM1 siRNA or Toll-like receptor 4 (TLR4) inhibitor induced low expression of TRAM1 in SC, which alleviated the pain response induced by CCI. The upregulation of p-NF-κB expression was reversed by TRAM1 siRNA in SC and DRG, and intrathecal injection of p-NF-κB inhibitor relieved neuropathic pain. All the data indicated that TRAM1 could take part in CCI-induced pain and might be a potential treatment for chronic neuropathic pain.     

The L2/HNK-1 Carbohydrate Epitope is Involved in the Preferential Outgrowth of Motor Neurons on Ventral Roots and Motor Nerves

Based on the observation that in adult mice the carbohydrate epitope L2/HNK-1 is detectable on Schwann cells in ventral spinal roots, but only scarcely in dorsal roots (Martini et al., Dev. Biol., 129, 330 - 338, 1988), the possibility was investigated that the carbohydrate is involved in the outgrowth of regenerating motor neuron axons on peripheral nerve substrates expressing the epitope. To monitor whether the L2 carbohydrate remains present during the time periods in which regenerating axons penetrate the denervated distal nerve stumps, the expression of L2 in motor and sensory branches of the femoral nerve was investigated in normal animals and after a crush lesion. During the first two postoperative weeks, L2 immunoreactivity remained high in the myelinating Schwann cells of the motor branch, whereas L2 immunoreactivity was virtually absent in the sensory branch. In a first experimental approach, cryosections of ventral and dorsal spinal roots and of motor and sensory nerves of adult rats and mice were used as substrates for neurite outgrowth. Neurites of motor neurons from chicken embryos were approximately 35% longer after 30 h of maintenance on ventral roots than on dorsal roots. Neurites from sensory neurons had the same length on dorsal as on ventral motors and were as long as neurites from motor neurons grown on dorsal roots. L2 antibodies reduced neurite outgrowth of motor neurons on ventral roots but not on dorsal roots. Neurite outgrowth of sensory neurons on both roots was not altered by the antibodies. Neurite outgrowth of motor neurons on a mixture of the extracellular matrix glycoprotein laminin and the L2 carbohydrate-carrying glycolipid was significantly higher than on the laminin substrate mixture with GD1b ganglioside or sulphatide. L2 antibodies reduced neurite outgrowth of motor neurons by 50% on the L2 glycolipid, but not on GD1b or sulphatide. These observations indicate that the L2 carbohydrate promotes neurite outgrowth of motor neurons in vitro and may thus contribute to the preferential reinnervation of motor nerves by regenerating motor axons in vivo.     

Synaptic Evoked Potentials from Regenerating Dorsal Root Axons within Fetal Spinal Cord Tissue Transplants

Previously injured dorsal roots were electrically stimulated to determine if regenerating sensory axons can form physiologically active synaptic contacts with neurons within fetal spinal cord tissue transplants. Dorsal rootlets, sectioned at their spinal cord entry zone, were apposed to intraspinal transplants of fetal spinal cord tissue grafted along each side of a nerve growth factor-treated nitrocellulose implant. Two to six months later, the rootlets were transected between the spinal cord and their respective ganglia and electrically stimulated. Evoked potentials were recorded from the dorsal surface of the transplant, but were absent from adjacent ipsilateral and contralateral spinal cord regions. A glass micropipette was advanced through the transplant and used to record intramedullary field potentials evoked by dorsal root stimulation. Maximal negative potentials occurred 400–700 μm below the dorsal surface of the transplant, shifting to positive potentials deeper into the transplant. Additionally, both spontaneous and electrically evoked single neuronal action potentials were observed along the microelectrode track. Evoked potentials were abolished following transection of the rootlets between the stimulation site and the transplant. Immunocytochemical evidence of the production of fos protein following electrical stimulation of the regenerated dorsal rootlets was demonstrated within transplant neurons and some ventrally located host neurons, providing an anatomical correlate to the electrophysiological recordings of synaptic activation. These results provide evidence of the structural and functional integration of regenerated sensory axons with both transplant and host neurons.     

N‐Syndecan and HB‐GAM (Heparin‐Binding Growth‐Associated Molecule) associate with early axonal tracts in the rat brain

Heparin-Binding Growth-Associated Molecule (HB-GAM)/pleiotrophin is an 18 kDa extracellular matrix- and cell-surface-associated protein shown to enhance neurite outgrowth of perinatal forebrain neurones in vitro. The heparan sulphate proteoglycan N-syndecan ( 1 ) has been isolated as a receptor/coreceptor for the HB-GAM. We have investigated, whether HB-GAM and N-syndecan could have a similar role in neurite outgrowth and axon guidance in early axonal tracts of brain. In the present study N-syndecan was found to be spatiotemporally associated with the developing axonal tracts already on embryonic day 9 in rat, as revealed by coexpression with class III β-tubulin, which is one of the earliest neuronal markers ( 2 ; 3 ). Later, N-syndecan and HB-GAM were detected in the first afferent serotonergic projections arising from the pontine raphe nuclei. The expression pattern of HB-GAM peaked in the developing rhombencephalon at embryonic stage (E) 13–14. At the same time, N-syndecan was expressed in the developing raphe neurones growing neurites towards the diencephalon along HB-GAM immunoreactive pathways. When rhombencephalic neurones were cultured on decreasing concentrations of substrate-bound HB-GAM, E13 neurones showed a significantly better neurite outgrowth response than E11, E16 or E18 neurones. The neurite outgrowth of raphe neurones in vitro was inhibited by adding soluble heparin or N-syndecan into the culture medium, whereas addition of chondroitin sulphate had no effect. In a simple pathway assay, E13 raphe neurones selectively preferred attaching and growing neurites on pathways containing HB-GAM as compared with regions containing either laminin or fibronectin alone. Our results suggest that HB-GAM may function as a developmentally regulated cue for rhombencephalic neurones that possess N-syndecan on their cell membrane.     

Vasoactive intestinal peptide influences neurite outgrowth in cultured rat spinal cord neurons.

Vasoactive intestinal peptide (VIP) is a neuropeptide which has been shown to exhibit a wide range of neurotrophic effects both in vivo and in vitro. For the purpose of clarifying the effect of VIP on spinal cord neurons, we studied the effect of VIP on neurite outgrowth of fetal rat ventral and dorsal portions of spinal cord in cultures. VIP-treated ventral spinal cord cultures (VSCC), compared with control VSCC, had a significant neurite outgrowth at 10(-8), 10(-6), and 10(-4) M. The effect was considered to be concentration dependent. Morphological changes of the dorsal spinal cord cultures (DSCC) remained unchanged by VIP treatment. Because of their close sequence homology with VIP, PHI-27 (peptide, histidylisoleucine amide) and secretin were also examined with the same experimental conditions as was VIP. Both PHI-27 and secretin had neurite promoting effects in VSCC at 10(-8) and 10(-6) M, respectively. However, there were no neurite promoting effects in DSCC in both of them at any concentrations. VIP had the most potent effect on neurite outgrowth in VSCC, followed by PHI-27, and secretin in their effectiveness concentrations. Our data showing VIP, PHI-27 and secretin have neurotrophic action on VSCC and suggest that a potential therapeutic use of VIP and its related peptides in treating diseases that involve degeneration and death of spinal motor neurons, such as motor neuropathy and amyotrophic lateral sclerosis.     

Regeneration into the Spinal Cord of Transected Dorsal Root Axons Is Promoted by Ensheathing Glia Transplants

The permissivity of adult olfactory bulb to the ingrowth of olfactory axons could be due to the unique properties of ensheathing glia. To test whether these glial cells could be used to promote axonal regeneration in a spontaneously nonregenerating system, we transplanted suspensions of pure ensheathing cells into a rhizotomized spinal cord segment. Ensheathing cells were purified away from other cell types by immunoaffinity, using anti-p75 nerve growth factor receptor. After laminectomy at the lower thoracic level, the spinal cord was exposed and one dorsal root (T10) was completely transected at the cord entry point. The root stump was microsurgically anastomosed to the cord and a suspension of ensheathing cells was transplanted in the spinal cord at the dorsal root entry zone. Three weeks after transplantation, numerous regenerating dorsal root axons were observed reentering the spinal cord. Ingrowth of dorsal root axons was observed using Dil and antibodies against calcitonin gene-related peptide and growth-associated protein. Primary sensory afferents invaded laminae 1, 2, and 3, grew through laminae 4 and 5, and reached the dorsal grey commissure and lamina 4 of the contralateral side. We did not observe regenerating axons within the ipsilateral ventral horn and dorsal column. Transplanted ensheathing cells reached the same laminae as axons. Neither ensheathing cells nor regenerating axons invaded those laminae they did not inervate under normal circumstances. In conclusion, the regeneration of injured dorsal root axons into the adult spinal cord was possible after ensheathing glia transplantation. The use of ensheathing cells as stimulators of axonal growth might be generalized to other central nervous system injuries.     

Fibroblast growth factor treatment produces differential effects on survival and neurite outgrowth from identified bulbospinal neurons in vitro

The in vivo application of appropriate trophic factors may enhance regeneration of bulbospinal projections after spinal cord injury. Currently, little is known about the sensitivities of specific bulbospinal neuron populations to the many identified trophic factors. We devised novel in vitro assays to study trophic effects on the survival and neurite outgrowth of identified bulbospinal neurons. Carbocyanine dye crystals implanted into the cervical spinal cord of embryonic day (E)5 chick embryos retrogradely labeled developing bulbospinal neurons. On E8, dissociated cultures containing labeled bulbospinal neurons were prepared. Fibroblast growth factor (FGF)-2 (but not FGF-1) promoted the survival of bulbospinal neurons. FGF receptor expression was widespread in the E8 brainstem, but not detected in young bulbospinal neurons, suggesting that nonneuronal cells mediated the FGF-stimulated survival response. Astrocytes synthesize a variety of trophic factors, and astrocyte-conditioned medium (ACM) also promoted the survival of bulbospinal neurons. As might be expected, FGF-2 function blocking antibodies did not suppress ACM-promoted survival, nor did an ELISA detect FGF-2 in ACM. This suggests that nonneuronal cells synthesize other factors in response to exogenous FGF-2 which promote the survival of bulbospinal neurons. Focusing on vestibulospinal neurons, dissociated (survival assay) or explant (neurite outgrowth assay) cultures were prepared. FGF-2 promoted both survival and neurite outgrowth of identified vestibulospinal neurons. Interestingly, FGF-1 promoted neurite outgrowth but not survival; the converse was true of FGF-9. Thus, differential effects of specific growth factors on survival or neurite outgrowth of bulbospinal neurons were distinguished.     

Immunohistochemical detection of estrogen receptor alpha in male rat spinal cord during development

The alpha subtype of the estrogen receptor (ERalpha) is present in nociceptive and parasympathetic regions of the adult rat spinal cord. The pattern of ERalpha expression in the rat spinal cord during development, however, is unknown. We used a polyclonal antibody (ER-21) to examine the expression of ERalpha in male rat lumbosacral spinal cords at embryonic day (E) 17, E21 (the day before birth), postnatal day (P) 1 (the day of birth), P8, P17, P21, and P36. At E17, ERalpha immunoreactivity (ERalpha-ir) was observed predominantly in ependymal cells. Perinatally, ERalpha-ir was also present in neurons in dorsal root ganglia and in fibers capping and within laminae I and II. By P8, ERalpha-ir was absent in ependymal cells, but ERalpha-ir fibers were dense in laminae I and II and in sympathetic and parasympathetic areas. ERalpha-ir was also present in neurons in the dorsal horns. To determine whether ERalpha-ir fibers in laminae I and II were processes of spinal neurons or primary afferents, dorsal rhizotomies were performed on P17 and P21 animals. Unilateral transection of the lumbosacral dorsal roots virtually eliminated ERalpha-ir fibers in the ipsilateral superficial laminae, demonstrating that the majority of ERalpha-ir fibers in these laminae were primary afferents. We show for the first time that ERalpha-ir is present in neurons and fibers of male prenatal and postnatal spinal cord. The presence of ERalpha in neuronal nuclei and processes may reflect diverse roles and novel mechanisms of action for 17 beta-estradiol in development of spinal sensory and autonomic circuitry.     

ROCK inhibition enhances neurite outgrowth in neural stem cells by upregulating YAP expressionin vitro

Spontaneous axonal regeneration of neurons does not occur after spinal cord injury because of inhibition by myelin and other inhibitory factors. Studies have demonstrated that blocking the Rho/Rho-kinase (ROCK) pathway can promote neurite outgrowth in spinal cord injury models. In the present study, we investigated neurite outgrowth and neuronal differentiation in neural stem cells from the mouse subventricular zone after inhibition of ROCK in vitro. Inhibition of ROCK with Y-27632 increased neurite length, enhanced neuronal differentiation, and upregulated the expression of two major signaling pathway effectors, phospho-Akt and phospho-mitogen-activated protein kinase, and the Hippo pathway effector YAP. These results suggest that inhibition of ROCK mediates neurite outgrowth in neural stem cells by activating the Hippo signaling pathway.     

Development of central projections of lumbosacral sensory neurons in the chick

The development of central projections of sensory neurons in lumbosacral dorsal root ganglia (DRGs) was examined by using horseradish peroxidase labeling techniques in chick embryos from stage 23 (E4) to stage 39 (E13). Our results show that primary afferents reach the spinal cord by stage 23. Afferent axons extend in the primordium of the dorsal funiculus for several segments rostral and caudal to their segment of entry for over 24 hours before invading the gray matter at stage 28 (E6). Sensory fibers grow into the vicinity of motoneuron dendrites by stage 32 (E7.5), about the time that reflexes and apparent monosynaptic EPSPs can first be elicited. Dense projections into the dorsal laminae of the spinal cord, presumably representing cutaneous afferents, appear somewhat later, at about stage 39 (E13), when the segmental projection pattern begins to resemble the mature pattern.     

Expression of c-Jun as a Response to Dorsal Root and Peripheral Nerve Section in Damaged and Adjacent Intact Primary Sensory Neurons in the Rat

It was previously shown that the immediate early gene, c-jun , was highly expressed over long periods, in both peripheral sensory and motor neurons following axon damage or block of axoplasmic transport. Here we have examined the question of whether the expression of c-Jun protein is related to axon injury per se or to the process of axon growth. We have examined dorsal root ganglion (DRG) cells subjected to different manipulations which are associated with varying degrees of regrowth, as follows: (i) after peripheral nerve section, where it appears that all damaged neurons make some regenerative effort. 1 – 24 days after sciatic nerve section and ligation most cells in L4/L5 DRG were c-Jun-positive; (ii) after section of the central processes of the DRG cells, which then showed a slow and limited regrowth of their axons towards, but not into, the spinal cord. This resulted in a variable, but significant, expression of c-Jun in a small number of DRG cells; (iii) in intact sensory neurons that were offered the opportunity to sprout into adjacent denervated peripheral tissue. The sciatic nerve was ligated and the response of cells in the L3 ganglia (many of which project to the saphenous nerve) was measured. A small but significant number of cells were c-Jun-positive; (iv) in intact sensory neurons that were offered the opportunity to sprout centrally into partialy denervated neuropil of the spinal cord. We examined neurons in the L3 DRG after rhizotomy of the adjacent L4/L5 dorsal roots. Previous work suggests that sensory neurons show at best a very limited growth under these conditions. No significant increase was seen in c-Jun expression in these cases. These results suggest that c-Jun expression is closely correlated with growth and regeneration, and not simply a consequence of neuronal injury.