Motheaten (me/me) mice deficient in SHP-1 are less susceptible to focal cerebral ischemia

We have demonstrated previously that the protein tyrosine phosphatase SHP-1 seems to play a role in glial development and is upregulated in non-dividing astrocytes after injury. The present study examines the effect of loss of SHP-1 on the CNS response to permanent focal ischemia. SHP-1 deficient (me/me) mice and wild-type littermates received a permanent middle cerebral artery occlusion (MCAO). At 1, 3, and 7 days after MCAO, infarct volume, neuronal survival and cell death, gliosis, and inflammatory cytokine levels were quantified. SHP-1 deficient me/me mice display smaller infarct volumes at 7 days post-MCAO, increased neuronal survival within the ischemic penumbra, and decreased numbers of cleaved caspase 3+ cells within the ischemic core compared with wild-type mice. In addition, me/me mice exhibit increases in GFAP+ reactive astrocytes, F4-80+ microglia, and a concomitant increase in the level of interleukin 12 (IL-12) over baseline compared with wild-type. Taken together, these results demonstrate that loss of SHP-1 results in greater healing of the infarct due to less apoptosis and more neuronal survival in the ischemic core and suggests that pharmacologic inactivation of SHP-1 may have potential therapeutic value in limiting CNS degeneration after ischemic stroke.     

Lipopolysaccharide-activated SHP-1-deficient motheaten microglia release increased nitric oxide, TNF-alpha, and IL-1beta

Accumulating evidence suggests a deleterious role for activated microglia in facilitating neuronal death by producing neurocytotoxic substances during injury, infection, or neurodegenerative diseases. After cochlear ablation, abnormal microglial activation accompanied by increased neuronal loss within the auditory brainstem occurs in motheaten (me/me) mice deficient in the protein tyrosine phosphatase SHP-1. To determine whether abnormally activated microglia contribute to neuronal death in me/me mice, primary microglial cultures from me/me and wild-type mouse cortices were stimulated by the bacterial endotoxin lipopolysaccharide (LPS) to evaluate the secretion of the neurotoxic mediators nitric oxide (NO), tumor necrosis factor-alpha (TNF-alpha), and interleukin-1beta (IL-1beta). Me/me microglia release significantly greater amounts of all three mediators compared with wild-type microglia. However, the increased release of these compounds in microglia lacking SHP-1 does not appear to occur through activation of extracellular signal-regulated kinase (ERK), p38 kinase subgroups of mitogen-activated protein (MAP) kinases, or increases in NF-kappaB-inducing kinase (NIK). These results suggest that abnormal microglial activation and release of neurotoxic compounds may potentiate neuronal death in deafferented cells and can thus potentiate neurodegeneration in the me/me brainstem. Our data also indicate that SHP-1 is engaged in signaling pathways in LPS-activated microglia, but not through regulation of the ERK and p38 MAP kinases.     

Tyrosine phosphatase SHP-1 immunoreactivity increases in a subset of astrocytes following deafferentation of the chicken auditory brainstem

Proliferation of astrocytes is a dramatic response of the central nervous system (CNS) to injury and disease. Such proliferation results in the formation of the neural/glial scar and the reconstitution of the glial limitans. However, not all astrocytes enter the proliferative cycle following injury, and for those that do, the period of cell division is limited. Little attention has focused on the events that regulate the duration and extent of astrocyte proliferation following damage, but clearly control mechanisms are in place as CNS injury does not result in the continuous astrocyte proliferation seen in glial tumorigenesis. Protein tyrosine phosphorylation has been implicated in both astrocyte proliferation and differentiation and plays an important role in the regulation of the cell cycle in a number of different systems. We have found a small subset of astrocytes in the chick auditory brainstem that are immunopositive for the protein tyrosine phosphatase SHP-1. SHP-1 appears to negatively regulate cellular division in the hematopoietic system and is involved in the mitogenic response to various growth factors. Following cochlea removal, there is a marked increase within the auditory brainstem nucleus, nucleus magnocellularis (NM), in both in the number of SHP-1-positive astrocytes and the length of their immunopositive fibers. Significantly, those animals showing the greatest increases in SHP-1 immunoreactivity do not exhibit large amounts of astrocyte proliferation. We hypothesize that the expression of SHP-1 plays a role in negatively regulating the mitotic behavior of astrocytes following deafferentation.     

Focal cerebral ischemia upregulates SHP-1 in reactive astrocytes in juvenile mice

The role of the tyrosine phosphatase SHP-1 in the hematopoietic system has been well studied; however, its role in the central nervous system (CNS) response to injury is not well understood. Previous studies in our laboratory have demonstrated increased immunoreactivity for SHP-1 in a subset of reactive astrocytes that do not appear to enter the cell cycle following deafferentation of the chicken auditory brainstem. In order to determine whether mammalian astrocytes also upregulate SHP-1 immunoreactivity following CNS injury, a mouse model of focal cerebral ischemia was utilized to study SHP-1 expression. The brains of 3-week-old mice were analyzed at four time points following permanent middle cerebral artery occlusion (MCAO): 1, 3, 7, and 14 days. Our results demonstrate consistent infarct volumes within surgical groups, and infarct volumes decrease as a function of time from 1 day (maximum infarct volume) to 14 days (minimum infarct volume) post-MCAO. In addition, SHP-1 protein levels are upregulated following cerebral ischemia and this increase peaks at 7 days post-MCAO. Analysis of confocal images further reveals that immunoreactivity for SHP-1 occurs predominantly in GFAP+ reactive astrocytes, although a small percentage of F4-80+ microglia are also double labeled for SHP-1 at early times post-MCAO. These SHP-1+ reactive astrocytes do not appear to enter the cell cycle (as defined by PCNA immunoreactivity), confirming our previous studies in the avian auditory brainstem. These results suggest that SHP-1 plays an important role in the regulation of glial activation and proliferation in the ischemic CNS.     

Expression and function of the protein tyrosine phosphatase SHP-1 in oligodendrocytes

Previous studies in this laboratory have shown that the SH-2 domain-containing protein tyrosine phosphatase SHP-1 is expressed in CNS glia and functions to modulate cytokine activities in these cells. The present study demonstrates that SHP-1 is expressed within multiple regions of the CNS in vivo, especially in white matter. Interestingly, we show that mice genetically lacking in SHP-1 (motheaten mice) in the CNS displayed dysmyelination. We therefore examined the expression of SHP-1 in the myelin-forming oligodendrocytes. Oligodendrocytes present in either mixed glial cultures or pure cultures expressed high levels of SHP-1 in the cytoplasm of cell bodies and processes. Oligodendrocytes isolated from motheaten mice did not express SHP-1. To test possible functions for SHP-1 in oligodendrocytes in controlling cytokine signaling, we compared the responsiveness of oligodendrocytes isolated from either motheaten or normal littermate mice with IL-6. IL-6 induced higher levels of STAT3 phosphorylation and STAT3-responsive c-fos gene expression in pure oligodendrocyte cultures of motheaten compared with normal littermate mice. These studies demonstrate that oligodendrocytes express SHP-1 and that SHP-1 functions to control IL-6 signaling. SHP-1 may therefore be a critical regulator of oligodendrocyte differentiation in response to IL-6 family cytokines. Further, these findings may relate to dysmyelination in mice lacking SHP-1.     

Inverse regulation of inducible nitric oxide synthase (iNOS) and arginase I by the protein tyrosine phosphatase SHP-1 in CNS glia

We have previously shown that the SH2 domain-containing protein tyrosine phosphatase SHP-1 plays a critical role in controlling virus infection in CNS glia in vivo and in vitro. The present study addressed whether increased virus replication in SHP-1-deficient glia in vitro may be a result of altered expression of inducible nitric oxide synthase (iNOS/NOS2). First, we observed a profound reduction in iNOS protein expression and production of nitric oxide (NO) in response to the viral mimic double-stranded RNA (dsRNA), despite the induction of high levels of iNOS mRNA, in SHP-1-deficient motheaten mouse compared to wild type littermate mouse glia. Because both iNOS expression and NO production are suppressed by multiple pathways involving arginase I activity, it was important that we observed abnormally high constitutive expression of arginase I in cultured glia of SHP-1-deficient compared to wild type mice. Further, both constitutive and IL-4/IL-10-induced expression of arginase I correlated with elevated STAT6 nuclear binding activity, decreased NO production, and increased virus replication in motheaten compared to wild type astrocytes. These findings provide the first evidence of an inverse relationship between NO and arginase I activity regulated by SHP-1 in CNS glia that is relevant to modulation of innate anti-viral responses. Thus, we propose that SHP-1 is a critical regulator of innate immunity to virus infections in CNS cells.     

Absence of brain-derived neurotrophic factor and trkB receptor immunoreactivity in glia of Alzheimer’s disease

Alterations in the neuronal expression of some neurotrophins have been shown in various neurodegenerative processes, particularly Alzheimer’s disease (AD). Glia may up-regulate neurotrophins and their high-affinity tyrosine kinase (trk) receptors in response to neural injury. In human immunodeficiency virus type 1 (HIV-1) encephalitis, activated microglia were shown to express brain-derived neurotrophic factor (BDNF), while reactive astrocytes expressed trkB receptor. This observation has suggested the existence of local neurotrophic regulation between different glial populations. To characterize the glial cellular distribution of BDNF and trkB receptor proteins in AD, we studied selected regions of postmortem brains from four AD and three age-matched control patients by double-immunofluorescence confocal microscopy. In both groups, BDNF immunoreactivity was distributed in neuronal perikarya and neuritic processes in the neocortex and hippocampus. No BDNF immunoreactivity was observed in microglia or astrocytes within and between senile plaques of AD. Catalytic trkB receptor immunoreactivity was present in neuronal perikarya in the neocortex and hippocampus. Reactive astrocytes and microglia were not immunoreactive for catalytic trkB. The absence of BDNF and trkB proteins in glia in AD patients is in contrast to the finding in patients with HIV-1 encephalitis. This difference suggests that glial expression of BDNF and trkB proteins may be characteristic of particular disease processes, rather than merely representing a stereotyped response to any type of neural injury. Received: 13 July 1998 / Revised, accepted: 25 February 1999     

Interleukin 15 expression in the CNS: blockade of its activity prevents glial activation after an inflammatory injury

Although reactive glia formation after neuronal degeneration or traumatic damage is one of the hallmarks of central nervous system (CNS) injury, we have little information on the signals that direct activation of resting glia. IL-15, a pro-inflammatory cytokine involved in regulating the response of T and B cells, may be also key for the regulation of early inflammatory events in the nervous system. IL-15 was expressed in the CNS, most abundantly in cerebellum and hippocampus, mainly in astrocytes and in some projection neurons. Using a rodent model of acute inflammatory injury [lipopolysaccharide (LPS) injection], we found enhanced expression of IL-15 in both reactive astroglia and microglia, soon after CNS injury. Blockade of IL-15 activity with an antibody to the cytokine, reversed activation of both glial types, suggesting that IL-15 has a major role in the generation of gliotic tissue and in the regulation of neuroimmune responses. Because IL-15 appears to modulate the inflammatory environment acutely generated after CNS injury, regulating IL-15 expression seems a clear antiinflammatory therapy to improve the outcome of neurodegenerative diseases and CNS trauma.     

Astroglia and microglia in cerebellar neuronal migration disturbances

Between the neuronal and glial cells there is a close relationship conditioning a tight morphological correlation and proper functional interplay. Disturbed interaction between glial and neuronal components leads to inappropriate neural circuits. The reflection of the failure of neural circuit organisation is the picture of morphological changes of neurons and glia. The appearance of microglia and astroglia was analysed in a defectively formed cellular network due to cerebellar neuronal migration disturbances. Focal disruption of neuron migration leads to their differentiation in an abnormal position manifested as heterotopias and cortical anomalies. Neurons that had lost their proper migratory way and heterotopically settled in the white matter were encircled by GFAP-positive astrocytes, with morphology appropriate for surrounding white matter. The microglial cells infiltrated the parenchyma within the heterotopic neurons playing a role in their elimination. In the cerebellar cortical malformations astrocytes were grouped near the Purkinje cells. In the minimal cortical dysplasia the increased number of astrocytes supported the neurons. Impaired morphological components of the glial-pial barrier were observed. In the massive cortical malformations a few degenerated astrocytes followed the disarranged Purkinje cells, while microglia and Bergmann glia fibres were not present. Absence of cells supporting and organizing the cerebellar cortex had an effect on loss of Purkinje cell shape, their disorientation and abnormal position. The appearance and localisation of the astroglia and microglia in the abnormal cerebellar circuitry due to migration disturbances is dependent on the pathomechanism of the anomalies.     

Effects of methylmercury on the secretion of pro-inflammatory cytokines from primary microglial cells and astrocytes

Glial cells, including oligodendrocytes, astrocytes and microglia are important to proper central nervous system (CNS) function. Deregulation or changes to CNS populations of astrocytes and microglia in particular are expected to play a role in many neurodegenerative diseases, including Parkinson's disease, amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD). Previous studies have reported methylmercury (MeHg) induced changes in glial cell function; however, the effects of MeHg on these cells remains poorly understood. This study aims to examine the effect of MeHg on the secretion of pro-inflammatory cytokines from microglia and astrocytes. The impact of the microglia/astrocyte ratio on cytokine secretion was also examined. Microglia and astrocytes were cultured from the brains of neo-natal BALB/C mice and dosed with MeHg (0-1 μM) and stimulated with PAM(3)CSK(4) (PAM(3)), a toll-like receptor (TLR) ligand. After this, the secretion of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α) and interleukin-1-beta (IL-1β) was measured by ELISA. MeHg reduced the secretion of IL-6 in a dose dependant manner but did not effect the secretion of TNF-α. No change in IL-1β was observed in any treatments, indicating that PAM(3) cannot induce the secretion of this cytokine from glial cells. Additionally, the ratio of microglia/astrocyte had an effect on the secretion of IL-6 but not TNF-α. These results indicate that MeHg can modify the response of glial cells and the interactions with astrocytes can affect the response of the microglia cells in culture. These results are significant in understanding the potential relationship with MeHg and neurodegenerative diseases and for the interpretation of results of future in vitro studies using monoculture.     

Astroglia disturbances during development of the central nervous system in fetuses with Down's syndrome

Down's syndrome (DS), caused by aneuploidy of chromosome 21, is the most common chromosomal disorder. The most significant symptom of this disorder is mental retardation. Neuropathological changes found in the DS central nervous system (CNS), such as reduced number of neurons, alteration of synapses and synaptic spines or delayed myelination have been widely described. But there are only a few studies of DS-related glia disturbances. A growing number of astroglia new functions have recently been described. In our study we compared the number of astrocytes and radial glial cells in the frontal lobe of DS fetuses at 18-20 weeks of gestation with that observed in age-matched controls. We found a substantially increased number of glial fibrillary acid protein (GFAP) positive cells in all age range samples of DS brains. We also noticed that in our study astrocytes in DS brains seem to be morphologically more mature than in controls of corresponding age. The same observation was made for radial glia. Taking into consideration the role played by astroglia during CNS development we believe that any change in their number, reduced or increased, can affect CNS development and lead to disturbances of both neurogenesis and synaptogenesis. A possible correlation between the increased number of astroglia and disturbances in CNS development is discussed.     

Role of activated astrocytes in neuronal damage: Potential links to HIV-1-associated dementia

HIV-1-associated dementia (HAD) is an important complication of HIV-1 infection. Reactive astrogliosis is a key pathological feature in HAD brains and in other central nervous system (CNS) diseases. Activated astroglia may play a critical role in CNS inflammatory diseases such as HAD. In order to test the hypothesis that activated astrocytes cause neuronal injury, we stimulated primary human fetal astrocytes with HAD-relevant pro-inflammatory cytokine IL-1beta. IL-1beta-activated astrocytes induced apoptosis and significant changes in metabolic activity in primary human neurons. An FITC-conjugated pan-caspase inhibitor peptide FITC-VAD-FMK was used for confirming caspase activation in neurons. IL-1beta activation enhanced the expression of death protein FasL in astrocytes, suggesting that FasL is one of the potential factors responsible for neurotoxicity observed in HAD and other CNS diseases involving glial inflammation. Our data presented here add to the developing picture of role of activated glia in HAD pathogenesis.     

Ganglioside GD3 in radial glia and astrocytes in situ in brains of young and adult mice

Ganglioside GD3 occurs in immature cells in the neuroectoderm. However, with regard to particular cellular locations of GD3, rat brain has received more attention than mouse brain. In brains from neonatal mice the most intense GD3 immunostaining appears to occur in structures that differ from those that immunostain the most intensely in brains from neonatal rats (Cammer and Zhang: J Histochem Cytochem 44: 143–149, 1996). In the present study epifluorescence and confocal microscopy were used for the purpose of identifying the types of GD3-immunopositive structures in brains of neonatal, 2-week-old, and adult mice. Vibratome sections from mouse brains were double immunostained for GD3 and respective markers for macrophages, microglia, and cells belonging to the oligodendrocyte lineage. Surprisingly, none of those marker antigens immunostained intensely in the same respective structures as GD3. The GD3-positive structures, however, did resemble protoplasmic astrocytes and radial glia, some with GD3-positive end-feet at the glia limitans; however, we did not rule out the possibility that there might be some GD3 on the surfaces of prooligodendroblasts. The scarcity of glial fibrillary acidic protein (GFAP)-positive cells in brains of neonatal mice made it impractical to look for GD3+/GFAP+ structures that might belong to the astrocyte lineage. The Mu subunit of glutathione-S-transferase (Mu) was shown to label radial glia and the few GFAP-positive cells in brains of neonatal mice. Subsequently, confocal microscopy showed Mu and GD3 to be colocalized in radial glia and protoplasmic astrocytes in the neonate. In brains from mice ≥2 weeks of age GD3 immunostaining was demonstrated in GFAP-positive astrocytes, including reactive astrocytes. Much of the GD3 appeared to occur at the tips of astrocyte processes. It is suggested that GD3 in radial glia and astrocytes may function as a ligand enabling recognition of those structures by neurons or as a precursor of more complex gangliosides in neurons. © 1996 Wiley-Liss, Inc.     

Radial glial cells are present in the velum medullare of adult monkeys

As a rule, in thin mammalian CNS tissues such as the median eminence and the retina, radial glia remains the dominating macroglia in adulthood, whereas in most other regions of the brain radial glial is substituted by multipolar macroglia i.e. astroglia. The Velum medullare is another thin CNS tissue but there are no reports on the dominating macroglia forms of this structure. Thus, Golgi-impregnated sections of adult monkey brains were studied for the presence of radial glial cells. Indeed, this structure was found to be transversed by many radial glial fibres terminating with pial endfeet whereas in adjacent thick brain tissues the glia limitans was formed by marginal astrocytes. It is concluded that fibrous radial glia may dominate in adult mammalian and even primate CNS tissues with a thickness of up to 1 mm.     

Stem cell therapy for central nerve system injuries： glial cells hold the key

Mammalian adult central nerve system （CNS） injuries are devastating because of the intrinsic difficulties for effective neuronal regeneration. The greatest problem to be overcome for CNS recovery is the poor regeneration of neurons and myelin-forming cells, oligodendrocytes. Endogenous neural progenitors and transplanted exogenous neuronal stem cells can be the source for neuronal regeneration. However, because of the harsh local microenvironment, they usually have very low efficacy for functional neural regeneration which cannot compensate for the loss of neurons and oligodendrocytes. Glial cells （including astrocytes, microglia, oligodendrocytes and NG2 glia） are the majority of cells in CNS that provide support and protection for neurons. Inside the local microenvironment, glial cells largely influence local and transplanted neural stem cells survival and fates. This review critically analyzes current finding of the roles of glial cells in CNS regeneration, and highlights strategies for regulating glial cells＇ behavior to create a permissive microenvironment for neuronal stem cells.     

NOD2 plays an important role in the inflammatory responses of microglia and astrocytes to bacterial CNS pathogens

While glial cells are recognized for their roles in maintaining neuronal function, there is growing appreciation that resident central nervous system (CNS) cells initiate and/or augment inflammation following trauma or infection. We have recently demonstrated that microglia and astrocytes constitutively express nucleotide-binding oligomerization domain-2 (NOD2), a member of the novel nucleotide-binding domain leucine-rich repeat region containing a family of proteins (NLR) that functions as an intracellular receptor for a minimal motif present in all bacterial peptidoglycans. In this study, we have confirmed the functional nature of NOD2 expression in astrocytes and microglia and begun to determine the relative contribution that this NLR makes in inflammatory CNS responses to clinically relevant bacterial pathogens. We demonstrate the increased association of NOD2 with its downstream effector molecule, Rip2 kinase, in primary cultures of murine microglia and astrocytes following exposure to bacterial antigens. We show that this cytosolic receptor underlies the ability of muramyl dipeptide to augment the production of inflammatory cytokines by glia following exposure to specific ligands for disparate Toll-like receptor homologues. In addition, we demonstrate that NOD2 is an important component in the in vitro inflammatory responses of resident glia to N. meningitidis and B. burgdorferi antigens. Finally, we have established that NOD2 is required, at least in part, for the astrogliosis, demyelination, behavioral changes, and elevated inflammatory cytokine levels observed following in vivo infection with these pathogens. As such, we have identified NOD2 as an important component in the generation of damaging CNS inflammation following bacterial infection.     

Novel insights into the glia limitans of the olfactory nervous system

Olfactory ensheathing cells (OECs) are often described as being present in both the peripheral and the central nervous systems (PNS and CNS). Furthermore, the olfactory nervous system glia limitans (the glial layer defining the PNS–CNS border) is considered unique as it consists of intermingling OECs and astrocytes. In contrast, the glia limitans of the rest of the nervous system consists solely of astrocytes which create a distinct barrier to Schwann cells (peripheral glia). The ability of OECs to interact with astrocytes is one reason why OECs are believed to be superior to Schwann cells for transplantation therapies to treat CNS injuries. We have used transgenic reporter mice in which glial cells express DsRed fluorescent protein to study the cellular constituents of the glia limitans. We found that the glia limitans layer of the olfactory nervous system is morphologically similar to elsewhere in the nervous system, with a similar low degree of intermingling between peripheral glia and astrocytes. We found that the astrocytic layer of the olfactory bulb is a distinct barrier to bacterial infection, suggesting that this layer constitutes the PNS–CNS immunological barrier. We also found that OECs interact with astrocytes in a similar fashion as Schwann cells in vitro. When cultured in three dimensions, however, there were subtle differences between OECs and Schwann cells in their interactions with astrocytes. We therefore suggest that glial fibrillary acidic protein–reactive astrocyte layer of the olfactory bulb constitutes the glia limitans of the olfactory nervous system and that OECs are primarily “PNS glia.”     

Deletion of oligodendrocyte Cx32 and astrocyte Cx43 causes white matter vacuolation, astrocyte loss and early mortality

CNS glia exhibit a variety of gap junctional interactions: between neighboring astrocytes, between neighboring oligodendrocytes, between astrocytes and oligodendrocytes, and as 'reflexive' structures between layers of myelin in oligodendrocytes. Together, these junctions are thought to form a network facilitating absorption and removal of extracellular K(+) released during neuronal activity. In mice, loss of the two major oligodendrocyte connexins causes severe demyelination and early mortality, while loss of the two major astrocyte connexins causes mild dysmyelination and sensorimotor impairment, suggesting that reflexive and/or oligo-oligo coupling may be more important for the maintenance of myelin than other forms. To further explore the functional relationships between glial connexins, we generated double knockout mice lacking one oligodendrocyte and one astrocyte connexin. Cx32-Cx43 dKO animals develop white matter vacuolation without obvious ultrastructural abnormalities in myelin. Progressive loss of astrocytes but not oligodendrocytes or microglia accompanies sensorimotor impairment, seizure activity and early mortality at around 16 weeks of age. Our data reveal an unexpected role for connexins in the survival of white matter astrocytes, requiring the expression of particular isoforms in both oligodendrocytes and astrocytes.     

Mice with GFAP-targeted loss of neurofibromin demonstrate increased axonal MET expression with aging

Neurofibromatosis 1 (NF1) is a common genetic disease that predisposes patients to peripheral nerve tumors and central nervous system (CNS) abnormalities including low-grade astrocytomas and cognitive disabilities. Using mice with glial fibrillary acidic protein (GFAP)-targeted Nf1 loss (Nf1(GFAP)CKO mice), we found that Nf1(-/-) astrocytes proliferate faster and are more invasive than wild-type astrocytes. In light of our previous finding that aberrant expression of the MET receptor tyrosine kinase contributes to the invasiveness of human NF1-associated malignant peripheral nerve sheath tumors, we sought to determine whether MET expression is aberrant in the brains of Nf1 mutant mice. We found that Nf1(-/-) astrocytes express slightly more MET than wild-type cells in vitro, but do not express elevated MET in situ. However, fiber tracts containing myelinated axons in the hippocampus, midbrain, cerebral cortex, and cerebellum express higher than normal levels of MET in older (> or =6 months) Nf1(GFAP)CKO mice. Both Nf1(GFAP)CKO and wild-type astrocytes induced MET expression in neurites of wild-type hippocampal neurons in vitro, suggesting that astrocyte-derived signals may induce MET in Nf1 mutant mice. Because the Nf1 gene product functions as a RAS GTPase, we examined MET expression in the brains of mice with GFAP-targeted constitutively active forms of RAS. MET was elevated in axonal fiber tracts in mice with active K-RAS but not H-RAS. Collectively, these data suggest that loss of Nf1 in either astrocytes or GFAP(+) neural progenitor cells results in increased axonal MET expression, which may contribute to the CNS abnormalities in children and adults with NF1.