Specific glial populations regulate hippocampal morphogenesis

The hippocampus plays an integral role in spatial navigation, learning and memory, and is a major site for adult neurogenesis. Critical to these functions is the proper organization of the hippocampus during development. Radial glia are known to regulate hippocampal formation, but their precise function in this process is yet to be defined. We find that in Nuclear Factor I b (Nfib)-deficient mice, a subpopulation of glia from the ammonic neuroepithelium of the hippocampus fail to develop. This results in severe morphological defects, including a failure of the hippocampal fissure, and subsequently the dentate gyrus, to form. As in wild-type mice, immature nestin-positive glia, which encompass all types of radial glia, populate the hippocampus in Nfib-deficient mice at embryonic day 15. However, these fail to mature into GLAST- and GFAP-positive glia, and the supragranular glial bundle is absent. In contrast, the fimbrial glial bundle forms, but alone is insufficient for proper hippocampal morphogenesis. Dentate granule neurons are present in the mutant hippocampus but their migration is aberrant, likely resulting from the lack of the complete radial glial scaffold usually provided by both glial bundles. These data demonstrate a role for Nfib in hippocampal fissure and dentate gyrus formation, and that distinct glial bundles are critical for correct hippocampal morphogenesis

NFIA controls telencephalic progenitor cell differentiation through repression of the Notch effector Hes1

The balance between self-renewal and differentiation of neural progenitor cells is an absolute requirement for the correct formation of the nervous system. Much is known about both the pathways involved in progenitor cell self-renewal, such as Notch signaling, and the expression of genes that initiate progenitor differentiation. However, whether these fundamental processes are mechanistically linked, and specifically how repression of progenitor self-renewal pathways occurs, is poorly understood. Nuclear factor I A (Nfia), a gene known to regulate spinal cord and neocortical development, has recently been implicated as acting downstream of Notch to initiate the expression of astrocyte-specific genes within the cortex. Here we demonstrate that, in addition to activating the expression of astrocyte-specific genes, Nfia also downregulates the activity of the Notch signaling pathway via repression of the key Notch effector Hes1. These data provide a significant conceptual advance in our understanding of neural progenitor differentiation, revealing that a single transcription factor can control both the activation of differentiation genes and the repression of the self-renewal genes, thereby acting as a pivotal regulator of the balance between progenitor and differentiated cell states

JNK/c-Jun Signaling and Peripheral Nerve Regeneration

The events associated with axonal injury in the peripheral nervous system (PNS) have been described in detail. However, the molecular mechanisms underlying the regenerative response, or neuronal cell death, following axonal injury are poorly understood. This thesis concerns one such molecular mechanism, JNK-mediated c-Jun activation, in peripheral sensory and sympathetic neurons. By immunohistochemistry, we demonstrate that the transcription factor c-Jun is rapidly activated by the mitogen activated protein kinase (MAPK) JNK, in both sensory and sympathetic neurons after axonal injury. Prevention of c-Jun activation with JNK inhibitors revealed that this activation is associated with survival and axonal outgrowth of developing sensory and sympathetic neurons. In adult sensory neurons, c-Jun activation is not required for survival, although it is required for axonal outgrowth. Additionally, c-Jun forms dimers with other transcription factors and it is plausible that the expression of these dimerization partners could regulate the physiological effect of c-Jun activation. We found that the activating transcription factor 3 (ATF3) was induced upon axonal injury in sensory and sympathetic neurons and that it colocalized with activated c-Jun. Consequently, we speculate that c-Jun/ATF3 dimers could be important for the regenerative response of peripheral neurons. Such dimers could promote a survival response in neurons under stress situations by inducing the expression of anti-apoptotic proteins such as heat shock protein 27 (Hsp27). We also made attempts to unravel the mechanism by which information of a distal axonal injury is conveyed to the cell body. We demonstrate that components of the JNK signaling pathway are axonally transported from the injury site to the cell body of dorsal root ganglia (DRG) sensory neurons, and also that this transport may contribute to the nuclear increase in c-Jun activity. Hence, we suggest that axonal transport of JNK signaling components could be involved in the transmission of injury information. Furthermore, since c-Jun activation may depend on deprivation of target-derived trophic factors, which is one consequence of axotomy, we analyzed the effect of nerve growth factor (NGF) on c-Jun activation in sensory and sympathetic neurons. NGF did not affect c-Jun activation in embryonic and adult sensory neurons, but it did affect c-Jun activation in neonatal sensory and sympathetic neurons. In conclusion, c-Jun activation seems to be a general neuronal response to peripheral nerve injury, and this response is associated with survival and regeneration

The role of p-c-Jun in survival and outgrowth of developing sensory neurons

c-jun activation has been implicated not only in neuronal apoptosis, but also in survival and regeneration. This Janus facet of c-Jun activation could be related to neuronal cell type or to the developmental stage of the neuron. We investigated c-Jun activation in E18 sensory neurons. Cultures of rat dorsal root ganglia neurons were maintained with or without the addition of nerve growth factor or the c-Jun N-terminal kinase inhibitor, (D)-JNKII. Few dorsal root ganglia neurons survived nerve growth factor deprivation, whereas neurons supplied with nerve growth factor survived and exhibited extensive axonal outgrowth. Activated c-Jun was present in the nuclei of neurons with regenerating axons, but not in apoptotic neurons. c-Jun N-terminal kinase inhibition reduced the number of p-c-Jun immunoreactive and regenerating neurons, and increased cell death. Thus, activation of c-Jun seems to be required for survival and regeneration of developing sensory neurons

Retrograde axonal transport of JNK signaling molecules influence injury induced nuclear changes in p-c-Jun and ATF3 in adult rat sensory neurons

In the present study, we investigated if the previously observed JNK-mediated activation of c-Jun and induction of ATF3 could be ascribed to axonal transport of JNK signaling components, or if axonal transport of the transcription factors themselves contributes to the nuclear changes in injured sensory neurons. We observed retrograde axonal transport of a number of JNK upstream kinases in ligated rat sciatic nerve. In these preparations, axonal transport of JNK/p-JNK, the JNK scaffolding protein JIP, and the transcription factors ATF3 and ATF2/p-ATF2 was also found. No or little retrograde transport of c-Jun and p-c-Jun was found, whereas an anterograde transport of Hsp27, a protein previously reported in the context of p-c-Jun and ATF3, was observed. In separate experiments, we found that in vitro inhibition of axonal transport or axonal inhibition of JNK reduced the number of p-c-Jun- and ATF3-positive neuronal nuclei. The results suggest that retrograde axonal transport of JNK signaling components contributes to the injury induced c-Jun phosphorylation and ATF3 induction

The Janus role of c-Jun: Cell death versus survival and regeneration of neonatal sympathetic and sensory neurons

We investigated the functional outcome of c-Jun activation in sympathetic and sensory neurons of neonatal rat superior cervical ganglion (SCG) and dorsal root ganglion (DRG), respectively. Distinctly different roles of c-Jun activation have been suggested for these two types of neurons. In dissociated sympathetic neurons, c-Jun has been demonstrated to promote apoptosis, whereas in sensory neurons it stimulates axonal outgrowth. In organ-cultured ganglia, we found that c-Jun was activated within 24 h of explantation in both types of neurons, and that the JNK inhibitor SP600125 could mitigate this response. In both types of neurons, c-Jun activation was also reduced by NGF treatment. Inhibition of c-Jun activation did not affect the viability of sympathetic neurons, whereas the number of apoptotic sensory neurons increased. Furthermore, inhibition of c-Jun reduced axonal outgrowth from both SCG and DRG. Thus, in organ culture, c-Jun activation may be required for axonal outgrowth and, at least in sensory neurons, it promotes survival. The role of ATF3, a neuronal marker of injury and a c-Jun dimerization partner, was also examined. We found an ATF3 induction in both SCG and DRG neurons, a response, which was reduced by JNK inhibition. The reduction of ATF3 upon JNK inhibition was much larger in DRG than in SCG, a result which might account for the higher number of apoptotic neurons in JNK inhibitor exposed DRG. Taken together, and contrary to our expectations, neonatal sympathetic and sensory neurons seem to respond to axonal injury similarly with respect to c-Jun activation, and in no case was this activation pro-apoptotic

Commissure formation in the mammalian forebrain

Commissural formation in the mammalian brain is highly organised and regulated both by the cell-autonomous expression of transcription factors, and by non-cell-autonomous mechanisms including the formation of midline glial structures and their expression of specific axon guidance molecules. These mechanisms channel axons into the correct path and enable the subsequent connection of specific brain areas to their appropriate targets. Several key findings have been made over the past two years, including the discovery of novel mechanisms of action that 'classical' guidance factors such as the Slits, Netrins, and their receptors have in axon guidance. Moreover, novel guidance factors such as members of the Writ family, and extracellular matrix components such as heparan sulphate proteoglycans, have been shown to be important for mammalian brain commissure formation. Additionally, there have been significant discoveries regarding the role of FGF signalling in the formation of midline glial structures. In this review, we discuss the most recent advances in the field that have contributed to our current understanding of commissural development in the telencephalon

Inhibition of c-Jun phosphorylation reduces axonal outgrowth of adult rat nodose ganglia and dorsal root ganglia sensory neurons.

The role of c-Jun activation for survival and regeneration of sensory neurons is unclear. Here we report that c-Jun N-terminal kinase (JNK)-mediated c-Jun activation is important for axonal outgrowth of sensory neurons in rat nodose and dorsal root ganglia (DRG). Peripheral severance of the vagus or the sciatic nerve resulted in a massive and rapid, but transient increase of the activated JNK (p-JNK) in neuronal nuclei, followed by c-Jun phosphorylation and activating transcription factor-3 (ATF3) induction. JNK inhibition by the selective JNK inhibitors SP600125 and (D)-JNKI1 did not affect neuronal survival in explanted or dissociated ganglia, but dramatically reduced axonal outgrowth, c-Jun activation, and ATF3 induction. Using retrograde labeling, we demonstrated that activated c-Jun (p-c-Jun) and ATF3 were associated with regenerative neurons. Taken together, our results suggest that JNK-mediated c-Jun activation is one of the first cell body reactions in response to nerve injury and that this activation and subsequent ATF3 induction are associated with axonal outgrowth

Chapter 28 future perspective in peripheral nerve reconstruction.

Nerve injuries induce severe disability and suffering for patients. Profound alterations in nerve trunks, neurons, and the central nervous system are induced rapidly after injury. This includes activation of intracellular signal transduction mechanisms aiming at the transfer of the cells into a regenerative state through the induction of the appropriate gene programs. The understanding of the neurobiological mechanisms that occur after injury can be used to design modern strategies for reconstruction after nerve injuries. Signal transduction mechanisms for instance may be targets for pharmacological intervention to stimulate nerve regeneration. Nerve injuries, particularly where there is a defect between the severed nerve trunks like in brachial plexus lesions, remain a challenge for the surgeon. Reconstruction of nerve injuries with a defect requires utilization of graft material, which can be of various designs. Application of autologous nerve grafts and use of nerve transfers are the most common clinical solutions to overcome problems with nerve defects. In this chapter we discuss the future perspective of nerve reconstruction with focus on signal transduction mechanisms and new avenues to bridge nerve defects using nanomodified graft surfaces

Ca2+ involvement in activation of extracellular-signalregulated- kinase 1/2 and m-calpain after axotomy of the sciatic nerve

Detailed mechanisms behind regeneration after nerve injury, in particular signal transduction and the fate of Schwann cells (SCs), are poorly understood. Here, we investigated axotomy-induced activation of extracellular- signal-regulated kinase-1/2 (ERK1/2; important for proliferation) and m-calpain in vitro, and the relation to Ca2+ deletion and Schwann cell proliferation and death after rat sciatic nerve axotomy. Nerve segments were cultured for up to 72 hours with and without ethylene glycol-bis(β-aminoethyl ether)- N,N,N’,N’-tetraacetic acid (EGTA). In some experiments, 5-bromo-2′-deoxyuridine (BrdU) was added during the last 24 hours to detect proliferating cells and propidium iodide (PI) was added at the last hour to detect dead and/or dying cells. Immunohistochemistry of sections of the cultured nerve segments was performed to label m-calpain and the phosphorylated and activated form of ERK1/2. The experiments revealed that immunoreactivity for p-ERK1/2 increased with time in organotypically cultured SCs. p-ERK1/2 and m-calpain were also observed in axons. A significant increase in the number of dead or dying SCs was observed in nerve segments cultured for 24 hours. When deprived of Ca2+, activation of axonal m-calpain was reduced, whereas p-ERK1/2 was increased in SCs. Ca2+ deprivation also significantly reduced the number of proliferating SCs, and instead increased the number of dead or dying SCs. Ca2+ seems to play an important role in activation of ERK1/2 in SCs and in SC survival and proliferation. In addition, extracellular Ca2+ levels are also required for m-calpain activation and up-regulation in axons. Thus, regulation of Ca2+ levels is likely to be a useful method to promote SC proliferation