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

Astrocytes in Alzheimer's Disease

The circuitry of the human brain is formed by neuronal networks embedded into astroglial syncytia. The astrocytes perform numerous functions, providing for the overall brain homeostasis, assisting in neurogenesis, determining the micro-architecture of the grey matter, and defending the brain through evolutionary conserved astrogliosis programs. Astroglial cells are engaged in neurological diseases by determining the progression and outcome of neuropathological process. Astrocytes are specifically involved in various neurodegenerative diseases, including Alzheimer’s disease, amyotrophic lateral sclerosis, Parkinson’s disease, and various forms of dementia. Recent evidence suggest that early stages of neurodegenerative processes are associated with atrophy of astroglia, which causes disruptions in synaptic connectivity, disbalance in neurotransmitter homeostasis, and neuronal death through increased excitotoxicity. At the later stages, astrocytes become activated and contribute to the neuroinflammatory component of neurodegeneration

A Combination of Ex vivo Diffusion MRI and Multiphoton to Study Microglia/Monocytes Alterations after Spinal Cord Injury

Central nervous system (CNS) injury has been observed to lead to microglia activation and monocytes infiltration at the lesion site. Ex vivo diffusion magnetic resonance imaging (diffusion MRI or DWI) allows detailed examination of CNS tissues, and recent advances in clearing procedures allow detailed imaging of fluorescent-labeled cells at high resolution. No study has yet combined ex vivo diffusion MRI and clearing procedures to establish a possible link between microglia/monocytes response and diffusion coefficient in the context of spinal cord injury (SCI). We carried out ex vivo MRI of the spinal cord at different time-points after spinal cord transection followed by tetrahydrofuran based clearing and examined the density and morphology of microglia/monocytes using two-photon microscopy. Quantitative analysis revealed an early marked increase in microglial/monocytes density that is associated with an increase in the extension of the lesion measured using diffusion MRI. Morphological examination of microglia/monocytes somata at the lesion site revealed a significant increase in their surface area and volume as early as 72 hours post-injury. Time-course analysis showed differential microglial/monocytes response rostral and caudal to the lesion site. Microglia/monocytes showed a decrease in reactivity over time caudal to the lesion site, but an increase was observed rostrally. Direct comparison of microglia/monocytes morphology, obtained through multiphoton, and the longitudinal apparent diffusion coefficient (ADC), measured with diffusion MRI, highlighted that axonal integrity does not correlate with the density of microglia/monocytes or their somata morphology. We emphasize that differential microglial/monocytes reactivity rostral and caudal to the lesion site may thus coincide, at least partially, with reported temporal differences in debris clearance. Our study demonstrates that the combination of ex vivo diffusion MRI and two-photon microscopy may be used to follow structural tissue alteration. Lesion extension coincides with microglia/monocytes density; however, a direct relationship between ADC and microglia/monocytes density and morphology was not observed. We highlighted a differential rostro-caudal microglia/monocytes reactivity that may correspond to a temporal difference in debris clearance and axonal integrity. Thus, potential therapeutic strategies targeting microglia/monocytes after SCI may need to be adjusted not only with the time after injury but also relative to the location to the lesion site

Correlation of in vivo and ex vivo H-1-MRI with histology in two severities of mouse spinal cord injury

Spinal cord injury (SCI) is a debilitating neuropathology with no effective treatment. Magnetic resonance imaging (MRI) technology is the only method used to assess the impact of an injury on the structure and function of the human spinal cord. Moreover, in pre-clinical SCI research, MRI is a non-invasive method with great translational potential since it provides relevant longitudinal assessment of anatomical and structural alterations induced by an injury. It is only recently that MRI techniques have been effectively used for the follow-up of SCI in rodents. However, the vast majority of these studies have been carried out on rats and when conducted in mice, the contusion injury model was predominantly chosen. Due to the remarkable potential of transgenic mice for studying the pathophysiology of SCI, we examined the use of both in and ex vivo 1H-MRI (9.4 T) in two severities of the mouse SCI (hemisection and over-hemisection) and documented their correlation with histological assessments. We demonstrated that a clear distinction between the two injury severities is possible using in and ex vivo 1H-MRI and that ex vivo MR images closely correlate with histology. Moreover, tissue modifications at a remote location from the lesion epicenter were identified by conventional ex vivo MRI analysis. Therefore, in vivo MRI has the potential to accurately identify in mice the progression of tissue alterations induced by SCI and is successfully implemented by ex vivo MRI examination. This combination of in and ex vivo MRI follow-up associated with histopathological assessment provides a valuable approach for further studies intended to evaluate therapeutic strategies on SCI

A Combination of Ex vivo Diffusion MRI and Multiphoton to Study Microglia/Monocytes Alterations after Spinal Cord Injury

Central nervous system (CNS) injury has been observed to lead to microglia activation and monocytes infiltration at the lesion site. Ex vivo diffusion magnetic resonance imaging (diffusion MRI or DWI) allows detailed examination of CNS tissues, and recent advances in clearing procedures allow detailed imaging of fluorescent-labeled cells at high resolution. No study has yet combined ex vivo diffusion MRI and clearing procedures to establish a possible link between microglia/monocytes response and diffusion coefficient in the context of spinal cord injury (SCI). We carried out ex vivo MRI of the spinal cord at different time-points after spinal cord transection followed by tetrahydrofuran based clearing and examined the density and morphology of microglia/monocytes using two-photon microscopy. Quantitative analysis revealed an early marked increase in microglial/monocytes density that is associated with an increase in the extension of the lesion measured using diffusion MRI. Morphological examination of microglia/monocytes somata at the lesion site revealed a significant increase in their surface area and volume as early as 72 hours post-injury. Time-course analysis showed differential microglial/monocytes response rostral and caudal to the lesion site. Microglia/monocytes showed a decrease in reactivity over time caudal to the lesion site, but an increase was observed rostrally. Direct comparison of microglia/monocytes morphology, obtained through multiphoton, and the longitudinal apparent diffusion coefficient (ADC), measured with diffusion MRI, highlighted that axonal integrity does not correlate with the density of microglia/monocytes or their somata morphology. We emphasize that differential microglial/monocytes reactivity rostral and caudal to the lesion site may thus coincide, at least partially, with reported temporal differences in debris clearance. Our study demonstrates that the combination of ex vivo diffusion MRI and two-photon microscopy may be used to follow structural tissue alteration. Lesion extension coincides with microglia/monocytes density; however, a direct relationship between ADC and microglia/monocytes density and morphology was not observed. We highlighted a differential rostro-caudal microglia/monocytes reactivity that may correspond to a temporal difference in debris clearance and axonal integrity. Thus, potential therapeutic strategies targeting microglia/monocytes after SCI may need to be adjusted not only with the time after injury but also relative to the location to the lesion site