C57BL/6 and Swiss Webster Mice Display Differences in Mobility, Gliosis, Microcavity Formation and Lesion Volume After Severe Spinal Cord Injury

Spinal cord injuries (SCI) are neuropathologies causing enormous physical and emotional anguish as well as irreversibly disabilities with great socio/economic burdens to our society. The availability of multiple mouse strains is important for studying the underlying pathophysiological response after SCI. Although strain differences have been shown to directly affect spontaneous functional recovery following incomplete SCI, its influence after complete lesion of the spinal cord is unclear. To study the influence of mouse strain on recovery after severe SCI, we first carried out behavioral analyses up to 6 weeks following complete transection of the spinal cord in mice with two different genetic backgrounds namely, C57BL/6 and Swiss Webster. Using immunohistochemistry, we then analyzed glial cell reactivity not only at different time-points after injury but also at different distances from the lesion epicenter. Behavioral assessments using CatWalk™ and open field analyses revealed increased mobility (measured using average speed) and differential forelimb gross sensory response in Swiss Webster compared to C57BL/6 mice after complete transection of the spinal cord. Comprehensive histological assessment revealed elevated microglia/macrophage reactivity and a moderate increase in astrogliosis in Swiss Webster that was associated with reduced microcavity formation and reduced lesion volume after spinal cord transection compared to C57BL/6 mice. Our results thus suggest that increased mobility correlates with enhanced gliosis and better tissue protection after complete transection of the spinal cord

Age-dependent decrease in glutamine synthetase expression in the hippocampal astroglia of the triple transgenic Alzheimer's disease mouse model : mechanism for deficient glutamatergic transmission?

Astrocytes are fundamental for brain homeostasis and the progression and outcome of many neuropathologies including Alzheimer's disease (AD). In the triple transgenic mouse model of AD (3xTg-AD) generalised hippocampal astroglia atrophy precedes a restricted and specific beta-amyloid (A beta) plaque-related astrogliosis. Astrocytes are critical for CNS glutamatergic transmission being the principal elements of glutamate homeostasis through maintaining its synthesis, uptake and turnover via glutamate-glutamine shuttle. Glutamine synthetase (GS), which is specifically expressed in astrocytes, forms glutamine by an ATP-dependent amination of glutamate. Here, we report changes in GS astrocytic expression in two major cognitive areas of the hippocampus (the dentate gyrus, DG and the CA1) in 3xTg-AD animals aged between 9 and 18 months. We found a significant reduction in Nv (number of cell/mm(3)) of GS immunoreactive (GS-IR) astrocytes starting from 12 months (28.59%) of age in the DG, and sustained at 18 months (31.65%). CA1 decrease of GS-positive astrocytes Nv (33.26%) occurs at 18 months. This Nv reduction of GSIR astrocytes is paralleled by a decrease in overall GS expression (determined by its optical density) that becomes significant at 18 months (21.61% and 19.68% in DG and CA1, respectively). GS-IR Nv changes are directly associated with the presence of A beta deposits showing a decrease of 47.92% as opposed to 23.47% in areas free of A beta. These changes in GS containing astrocytes and GS-immunoreactivity indicate AD-related impairments of glutamate homeostatic system, at the advanced and late stages of the disease, which may affect the efficacy of glutamatergic transmission in the diseased brain that may contribute to the cognitive deficiency.The present study was supported by Alzheimer's Research Trust Programme Grant (ART/PG2004A/1) to JJR and AV. Grant Agency of the Czech Republic (GACR 309/09/1696 and GACR 304/11/0184) to JJR and (GACR 305/08/1381; GACR 305/08/1384) to AV. The Spanish Government, Plan Nacional de I+D+I 2008-2011 and ISCIII-Subdireccion General de Evaluacion y Fomento de la investigacion (PI10/02738) to JJR and AV and the Government of the Basque Country grant (AE-2010-1-28; AEGV10/16) to JJR. The authors would also like to thank BBSRC for the Ph.D. studentship to H.N. Noristani

Microglia Responses in Acute and Chronic Neurological Diseases: What Microglia-Specific Transcriptomic Studies Taught (and did Not Teach) Us

Over the last decade, microglia have been acknowledged to be key players in central nervous system (CNS) under both physiological and pathological conditions. They constantly survey the CNS environment and as immune cells, in pathological contexts, they provide the first host defense and orchestrate the immune response. It is well recognized that under pathological conditions microglia have both sequential and simultaneous, beneficial and detrimental effects. Cell-specific transcriptomics recently became popular in Neuroscience field allowing concurrent monitoring of the expression of numerous genes in a given cell population. Moreover, by comparing two or more conditions, these approaches permit to unbiasedly identify deregulated genes and pathways. A growing number of studies have thus investigated microglial transcriptome remodeling over the course of neuropathological conditions and highlighted the molecular diversity of microglial response to different diseases. In the present work, we restrict our review to microglia obtained directly from in vivo samples and not cell culture, and to studies using whole-genome strategies. We first critically review the different methods developed to decipher microglia transcriptome. In particular, we compare advantages and drawbacks of flow cytometry and laser microdissection to isolate pure microglia population as well as identification of deregulated microglial genes obtained via RNA sequencing (RNA-Seq) vs. microarrays approaches. Second, we summarize insights obtained from microglia transcriptomes in traumatic brain and spinal cord injuries, pain and more chronic neurological conditions including Amyotrophic lateral sclerosis (ALS), Alzheimer disease (AD) and Multiple sclerosis (MS). Transcriptomic responses of microglia in other non-neurodegenerative CNS disorders such as gliomas and sepsis are also addressed. Third, we present a comparison of the most activated pathways in each neuropathological condition using Gene ontology (GO) classification and highlight the diversity of microglia response to insults focusing on their pro- and anti-inflammatory signatures. Finally, we discuss the potential of the latest technological advances, in particular, single cell RNA-Seq to unravel the individual microglial response diversity in neuropathological contexts

Neuronatin Promotes Neural Lineage in ESCs via Ca2+ Signaling

Neural induction is the first step in the formation of the vertebrate central nervous system. The emerging consensus of the mechanisms underling neural induction is the combined influences from inhibiting bone morphogenetic protein (BMP) signaling and activating fibroblast growth factor (FGF)/Erk signaling, which act extrinsically via either autocrine or paracrine fashions. However, do intrinsic forces (cues) exist and do they play decisive roles in neural induction? These questions remain to be answered. Here, we have identified a novel neural initiator, neuronatin (Nnat), which acts as an intrinsic factor to promote neural fate in mammals and Xenopus. ESCs lacking this intrinsic factor fail to undergo neural induction despite the inhibition of the BMP pathway. We show that Nnat initiates neural induction in ESCs through increasing intracellular Ca2+ ([Ca2+]i) by antagonizing Ca2+-ATPase isoform 2 (sarco/endoplasmic reticulum Ca2+-ATPase isoform 2) in the endoplasmic reticulum, which in turn increases the phosphorylation of Erk1/2 and inhibits the BMP4 pathway and leads to neural induction in conjunction with FGF/Erk pathway. STEM CELLS 2010;28:1950–196

Serotonergic Mechanisms in Spinal Cord Injury

International audienc

Astrocyte-to-neuron conversion induced by spinal cord injury

International audienceSpinal cord injury (SCI) triggers pronounced astrocyte reactivity (astrogliosis) including astroglial proliferation and migration toward the injury site participating to the formation of a glial scar. Since the mid-20 th century, SCI-induced astrogliosis was mainly regarded as detrimental for successful axonal regeneration. However, more recent studies have shown astrogliosis as a multifactorial phenomenon involving specific morphological, molecular and functional alterations in astrocytes that can also exert beneficial effects [1, 2]. It was suggested, although not proven, that SCI-induced astrogliosis depends on multiple factors such as time after lesion, injury severity and distance to the lesion site. In a recent study we had attempted to uncover the molecular involvement of astrocytes after SCI by studying their transcriptomic alterations at different stages after moderate and severe lesions [3]. Aldehyde dehydrogenase 1 family member L1 (Aldh1l1) is a pan-astrocytic marker, hence using the Aldh1l1-EGFP transgenic mice, combined with fluorescence-activated cell sorting (FACS), we isolated pure astrocyte population at different stages following SCI. Choosing lateral hemisection and complete section of the spinal cord, as moderate and severe injury models, we investigated astrocytic response at 1 and 2 weeks after lesion. We subsequently carried out astrocyte-specific RNA-sequencing and pathway analyses to unveil the molecular signature of injuries-induced astrogliosis. Our transcriptomic analyses demonstrated a dual astrocytic response depending on time post-injury and lesion severity. Following moderate SCI, astrocytes displayed a protective role and showed no changes (1 week) and even down-regulated (2 weeks) expression of transcripts involved in immune response. On the other hand, astrocytes response after severe SCI seems to be detrimental by an upsurge expression of inflammatory genes (1 week) and prevention of extracellular re-modeling (2 weeks) (3). These are the first concrete evidence of a heterogeneous astrocytic response that is driven not only by lesion severity but also time after injury (Figure 1). In parallel, using pathway analyses, we also identified in astrocytes the induction of the neural stem cell lineage and the over-expression of the neuronal progenitor gene βIII-tubulin (Tubb3, also known as TUJ1). We confirmed βIII-tubulin protein expression at tissue level using immunohistochemistry and at single cell level using FACS analyses. The sub-population of astrocytes that express βIII-tubulin was only found within 750µm distance to the lesion epicenter. Astrocytes co-expressing βIII-tubulin, also displayed alterations in their morphology from typical stellate shape to classical neuronal progenitor cells with bipolar or multipolar processes. Given that SCI induces astrocytic proliferation, we injected BrdU in Aldh1l1-EGFP mice after injury to determine the origin of eGFP/βIII-tubulin co-expressing cells. BrdU incorporation was observed into newly formed astrocytes but not in eGFP/βIII-tubulin-expressing astrocytes. This suggests that it is the resident mature astrocytes, rather than newly formed astrocytes, that undergo transdifferentiation towards neuronal lineage (Figure 1). Time-dependent analyses revealed that astrocytic conversion towards neuronal lineage starts as early as 72 hours, peaking between 1-2 weeks and continues to a lower degree up to 6 weeks after both moderate and severe SCI. Further immunostaining, using mature neuronal markers, showed that transdifferentiating astrocytes eventually express GABAergic, but not glutamatergic, markers. Moreover, we identified the fibroblast growth factor receptor 4 (Fgfr4) as a potential player responsible for SCI-induced astrocytic transdifferentiation towards neuronal lineage. Fgfr4 indeed promotes embryonic stem cell differentiation towards neuronal lineage [4] and showed pronounced over-expression from 72 hours following lesion at both RNA and protein level. Although other recent studies had shown limited astrocytes conversion towards neuronal lineage upon enforced expression of neurogenic factors, none had Editorial Figure 1: Schematic cartoon displaying summary of astrocytic responses following SC

Microglia Responses in Acute and Chronic Neurological Diseases: What Microglia-Specific Transcriptomic Studies Taught (and did Not Teach) Us

International audienceOver the last decade, microglia have been acknowledged to be key players in central nervous system (CNS) under both physiological and pathological conditions. They constantly survey the CNS environment and as immune cells, in pathological contexts, they provide the first host defense and orchestrate the immune response. It is well recognized that under pathological conditions microglia have both sequential and simultaneous, beneficial and detrimental effects. Cell-specific transcriptomics recently became popular in Neuroscience field allowing concurrent monitoring of the expression of numerous genes in a given cell population. Moreover, by comparing two or more conditions, these approaches permit to unbiasedly identify deregulated genes and pathways. A growing number of studies have thus investigated microglial transcriptome remodeling over the course of neuropathological conditions and highlighted the molecular diversity of microglial response to different diseases. In the present work, we restrict our review to microglia obtained directly from in vivo samples and not cell culture, and to studies using whole-genome strategies. We first critically review the different methods developed to decipher microglia transcriptome. In particular, we compare advantages and drawbacks of flow cytometry and laser microdissection to isolate pure microglia population as well as identification of deregulated microglial genes obtained via RNA sequencing (RNA-Seq) vs. microarrays approaches. Second, we summarize insights obtained from microglia transcriptomes in traumatic brain and spinal cord injuries, pain and more chronic neurological conditions including Amyotrophic lateral sclerosis (ALS), Alzheimer disease (AD) and Multiple sclerosis (MS). Transcriptomic responses of microglia in other non-neurodegenerative CNS disorders such as gliomas and sepsis are also addressed. Third, we present a comparison of the most activated pathways in each neuropathological condition using Gene ontology (GO) classification and highlight the diversity of microglia response to insults focusing on their pro- and anti-inflammatory signatures. Finally, we discuss the potential of the latest technological advances, in particular, single cell RNA-Seq to unravel the individual microglial response diversity in neuropathological contexts