Author Correction: Lipid-droplet-accumulating microglia represent a dysfunctional and proinflammatory state in the aging brain.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Author Correction: Lipid-droplet-accumulating microglia represent a dysfunctional and proinflammatory state in the aging brain.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The Leukotriene Receptor Antagonist Montelukast Reduces Alpha-Synuclein Load and Restores Memory in an Animal Model of Dementia with Lewy Bodies.

Dementia with Lewy bodies (DLB) represents a huge medical need as it accounts for up to 30% of all dementia cases, and there is no cure available. The underyling spectrum of pathology is complex and creates a challenge for targeted molecular therapies. We here tested the hypothesis that leukotrienes are involved in the pathology of DLB and that blocking leukotrienes through Montelukast, a leukotriene receptor antagonist and approved anti-asthmatic drug, might alleviate pathology and restore cognitive functions. Expression of 5-lipoxygenase, the rate-limiting enzyme for leukotriene production, was indeed elevated in brains with DLB. Treatment of cognitively deficient human alpha-synuclein overexpressing transgenic mice with Montelukast restored memory. Montelukast treatment resulted in modulation of beclin-1 expression, a marker for autophagy, and in a reduction in the human alpha-synulcein load in the transgenic mice. Reducing the protein aggregation load in neurodegenerative diseases might be a novel model of action of Montelukast. Moreover, this work presents leukotriene signaling as a potential drug target for DLB and shows that Montelukast might be a promising drug candidate for future DLB therapy development.

Lipid-droplet-accumulating microglia represent a dysfunctional and proinflammatory state in the aging brain.

Microglia become progressively activated and seemingly dysfunctional with age, and genetic studies have linked these cells to the pathogenesis of a growing number of neurodegenerative diseases. Here we report a striking buildup of lipid droplets in microglia with aging in mouse and human brains. These cells, which we call 'lipid-droplet-accumulating microglia' (LDAM), are defective in phagocytosis, produce high levels of reactive oxygen species and secrete proinflammatory cytokines. RNA-sequencing analysis of LDAM revealed a transcriptional profile driven by innate inflammation that is distinct from previously reported microglial states. An unbiased CRISPR-Cas9 screen identified genetic modifiers of lipid droplet formation; surprisingly, variants of several of these genes, including progranulin (GRN), are causes of autosomal-dominant forms of human neurodegenerative diseases. We therefore propose that LDAM contribute to age-related and genetic forms of neurodegeneration.

Developmental Heterogeneity of Microglia and Brain Myeloid Cells Revealed by Deep Single-Cell RNA Sequencing.

Microglia are increasingly recognized for their major contributions during brain development and neurodegenerative disease. It is currently unknown whether these functions are carried out by subsets of microglia during different stages of development and adulthood or within specific brain regions. Here, we performed deep single-cell RNA sequencing (scRNA-seq) of microglia and related myeloid cells sorted from various regions of embryonic, early postnatal, and adult mouse brains. We found that the majority of adult microglia expressing homeostatic genes are remarkably similar in transcriptomes, regardless of brain region. By contrast, early postnatal microglia are more heterogeneous. We discovered a proliferative-region-associated microglia (PAM) subset, mainly found in developing white matter, that shares a characteristic gene signature with degenerative disease-associated microglia (DAM). Such PAM have amoeboid morphology, are metabolically active, and phagocytose newly formed oligodendrocytes. This scRNA-seq atlas will be a valuable resource for dissecting innate immune functions in health and disease.

The leukotriene signaling pathway: a druggable target in Alzheimer's disease.

The underlying pathology of Alzheimer's disease (AD) is complex and includes, besides amyloid beta (Aß) plaque depositions and neurofibrillary tangles, brain atrophy and neurodegeneration, neuroinflammation, impaired neurogenesis, vascular and blood-brain barrier (BBB) disruptions, neurotransmitter disbalances, and others. Here, we hypothesize that such complex pathologies can only be targeted efficiently through pleiotropic approaches. One interesting drug target is the leukotriene pathway, which mediates various aspects of AD pathology. Approaching this pathway at different levels with genetic and pharmacological tools demonstrated beneficial outcomes in several in vivo studies using different mouse models of AD. Here, we review the current literature on the leukotriene signaling pathway as a target for drug development in AD.

Doublecortin expression in CD8+ T-cells and microglia at sites of amyloid-ß plaques: A potential role in shaping plaque pathology?

INTRODUCTION: One characteristic of Alzheimer's disease is the formation of amyloid-ß plaques, which are typically linked to neuroinflammation and surrounded by inflammatory cells such as microglia and infiltrating immune cells. METHODS: Here, we describe nonneurogenic doublecortin (DCX) positive cells, DCX being generally used as a marker for young immature neurons, at sites of amyloid-ß plaques in various transgenic amyloid mouse models and in human brains with plaque pathology. RESULTS: The plaque-associated DCX+ cells were not of neurogenic identity, instead most of them showed coexpression with markers for microglia (ionized calcium-binding adapter molecule 1) and for phagocytosis (CD68 and TREM2). Another subpopulation of plaque-associated DCX+ cells was negative for ionized calcium-binding adapter molecule 1 but was highly positive for the pan-leukocyte marker CD45. These hematopoietic cells were identified as CD3-and CD8-positive and CD4-negative T-cells. DISCUSSION: Peculiarly, the DCX+/ionized calcium-binding adapter molecule 1+ microglia and DCX+/CD8+ T-cells were closely attached, suggesting that these two cell types are tightly interacting and that this interaction might shape plaque pathology.

Motor deficits following dorsal corticospinal tract transection in rats: voluntary versus skilled locomotion readouts.

Following spinal cord injury, severe deficits result from damages to ascending and descending tracts, such as the corticospinal tract (CST) which is highly relevant for the motor execution in humans. Unfortunately, no curative treatment is available and intensive efforts are deployed in animal models, such as the CST transection model, to identify interventions providing functional regeneration after spinal cord injury. The CatWalk XT is a system for multi-parameter gait analysis of voluntary locomotion. In this study, the performance of the CatWalk XT for monitoring of functional deficits associated with dorsal CST lesion in rats was compared to skilled locomotion tests. Motor deficits associated with dorsal CST transection could be reliably monitored over seven weeks based on skilled locomotion testing, i.e. Horizontal Ladder Walk and Grid Walk. The collateral lesion to the overlaying gracile and cuneate funiculi occurring during dorsal CST transection resulted in slight hyposensitivity and proprioceptive deficit, which likely contributed to the lowered performance in skilled locomotion. In contrast, parameters of voluntary locomotion were not significantly affected by dorsal CST transection. Finally, an abnormal adduction reflex was detected immediately after lesion of the CST and could be conveniently used to confirm successful CST lesion in rats of experimental groups. The functional relevance of the dorsal CST in locomotion of rats is not as prominent as compared to in humans and thus challenging the motor execution is mandatory to reliably investigate CST function. A detailed analysis of voluntary walking using the CatWalk XT is not adequate to detect deficits following dorsal CST lesion in rats.

Tamoxifen Activation of Cre-Recombinase Has No Persisting Effects on Adult Neurogenesis or Learning and Anxiety.

Adult neurogenesis is a tightly regulated process continuously taking place in the central nervous system of most mammalian species. In neuroscience research, transgenic animals bearing the tamoxifen-inducible CreERT2-Lox system are widely used. In this study, we made use of a Nestin-CreERT2/R26R-YFP transgenic mouse model in which the CreERT2 activates the expression of YFP in multipotent neural stem cells upon tamoxifen application. Humoral factors, such as the levels of estrogens, have been reported to affect the hippocampal neurogenesis. The application of tamoxifen, a mixed agonist/antagonist of the estrogen receptor that permeates the blood-brain-barrier, could thus influence adult neurogenesis. Although the functions of adult neurogenesis are yet to be fully deciphered, a reciprocal interaction between rates of neurogenesis on the one hand and learning and mood regulation on the other hand, has been suggested. The impact of tamoxifen on neurogenesis and behavior was therefore addressed following five daily applications according to the open field test, the elevated plus maze, and Morris water maze. In addition, the impact of short-term tamoxifen application on progenitor cell proliferation, morphology, and fate in the neurogenic niche of the dentate gyrus were investigated. Finally, the influence of the route of administration (oral vs. intra-peritoneal) and gender-specific response were scrutinized. The sub-acute analysis did neither reveal significant differences in behavior, such as voluntary motor activity, anxiety behavior, and spatial learning, nor in cell proliferation, cell survival, dendritic arborization or maturation rate within the dentate gyrus between saline solution-, corn oil-, and tamoxifen-treated groups. Finally, neither the route of application, nor the gender of treated mice influenced the response to tamoxifen. We conclude that short tamoxifen treatments used to activate the CreERT2 system in transgenic mouse models does not have a measurable impact on adult neurogenesis or the here tested behavior, and is therefore appropriate for most studies in the field.

Nontraumatic spinal cord injury at the neurological intensive care unit: spectrum, causes of admission and predictors of mortality.

OBJECTIVE: Nontraumatic spinal cord injuries (NTSCIs) form a heterogeneous group of diseases, which may evolve into a life-threatening condition. We sought to characterize spectrum, causes of admission and predictors of death in patients with NTSCI treated at the neurological intensive care unit (NICU). METHODS: We performed a retrospective observational analysis of NTSCI cases treated at a tertiary care center between 2001 and 2013. Among the 3937 NICU admissions were 93 patients with NTSCI (2.4%). Using multivariate logistic regression analysis, we examined predictors of mortality including demographics, etiology, reasons for admission and GCS/SAPS (Glasgow Coma Scale/Simplified Acute Physiology Score) scores. RESULTS: Infectious and inflammatory/autoimmune causes made up 50% of the NTSCI cases. The most common reasons for NICU admission were rapidly progressing paresis (49.5%) and abundance of respiratory insufficiency (26.9%). The mortality rate was 22.6% and 2.5-fold higher than in the cohort of all other patients treated at the NICU. Respiratory insufficiency as the reason for NICU admission [odds ratio (OR) 4.97, 95% confidence interval (CI) 1.38-17.9; p < 0.01], high initial SAPS scores (OR 1.04; 95% CI 1.003-1.08; p = 0.04), and the development of acute kidney injury throughout the stay (OR 7.25, 1.9-27.5; p = 0.004) were independent risk factors for NICU death. CONCLUSIONS: Patients with NTSCI account for a subset of patients admitted to the NICU and are at risk for adverse outcome. A better understanding of predisposing conditions and further knowledge of management of critically ill patients with NTSCI is mandatory.

Structural and functional rejuvenation of the aged brain by an approved anti-asthmatic drug.

As human life expectancy has improved rapidly in industrialized societies, age-related cognitive impairment presents an increasing challenge. Targeting histopathological processes that correlate with age-related cognitive declines, such as neuroinflammation, low levels of neurogenesis, disrupted blood-brain barrier and altered neuronal activity, might lead to structural and functional rejuvenation of the aged brain. Here we show that a 6-week treatment of young (4 months) and old (20 months) rats with montelukast, a marketed anti-asthmatic drug antagonizing leukotriene receptors, reduces neuroinflammation, elevates hippocampal neurogenesis and improves learning and memory in old animals. By using gene knockdown and knockout approaches, we demonstrate that the effect is mediated through inhibition of the GPR17 receptor. This work illustrates that inhibition of leukotriene receptor signalling might represent a safe and druggable target to restore cognitive functions in old individuals and paves the way for future clinical translation of leukotriene receptor inhibition for the treatment of dementias.

The L-type calcium channel Cav1.3 is required for proper hippocampal neurogenesis and cognitive functions.

L-type voltage gated Ca(2+) channels (LTCCs) are widely expressed within different brain regions including the hippocampus. The isoforms Cav1.2 and Cav1.3 have been shown to be involved in hippocampus-dependent learning and memory, cognitive functions that require proper hippocampal neurogenesis. In vitro, functional LTCCs are expressed on neuronal progenitor cells, where they promote neuronal differentiation. Expression of LTCCs on neural stem and progenitor cells within the neurogenic regions in the adult brain in vivo has not been examined so far, and a contribution of the individual isoforms Cav1.2 and Cav1.3 to adult neurogenesis remained to be clarified. To reveal the role of these channels we first evaluated the expression patterns of Cav1.2 and Cav1.3 in the hippocampal dentate gyrus and the subventricular zone (SVZ) in adult (2- and 3-month old) and middle-aged (15-month old) mice on mRNA and protein levels. We performed immunohistological analysis of hippocampal neurogenesis in adult and middle-aged Cav1.3(-/-) mice and finally addressed the importance of Cav1.3 for hippocampal function by evaluating spatial memory and depression-like behavior in adult Cav1.3(-/-) mice. Our results showed Cav1.2 and Cav1.3 expression at different stages of neuronal differentiation. While Cav1.2 was primarily restricted to mature NeuN(+) granular neurons, Cav1.3 was expressed in Nestin(+) neural stem cells and in mature NeuN(+) granular neurons. Adult and middle-aged Cav1.3(-/-) mice showed severe impairments in dentate gyrus neurogenesis, with significantly smaller dentate gyrus volume, reduced survival of newly generated cells, and reduced neuronal differentiation. Further, Cav1.3(-/-) mice showed impairment in the hippocampus dependent object location memory test, implicating Cav1.3 as an essential element for hippocampus-associated cognitive functions. Thus, modulation of LTCC activities may have a crucial impact on neurogenic responses and cognition, which should be considered for future therapeutic administration of LTCCs modulators.

Hippocampal neurogenesis and antidepressive therapy: shocking relations.

Speculations on the involvement of hippocampal neurogenesis, a form of neuronal plasticity, in the aetiology of depression and the mode of action of antidepressive therapies, started to arise more than a decade ago. But still, conclusive evidence that adult neurogenesis contributes to antidepressive effects of pharmacological and physical therapies has not been generated yet. This review revisits recent findings on the close relation between the mode(s) of action of electroconvulsive therapy (ECT), a powerful intervention used as second-line treatment of major depression disorders, and the neurogenic response to ECT. Following application of electroconvulsive shocks, intricate interactions between neurogenesis, angiogenesis, and microglia activation, the hypothalamic-pituitary-adrenal axis and the secretion of neurotrophic factors have been documented. Furthermore, considering the fact that neurogenesis strongly diminishes along aging, we investigated the response to electroconvulsive shocks in young as well as in aged cohorts of mice.

TGF-beta signalling in the adult neurogenic niche promotes stem cell quiescence as well as generation of new neurons.

Members of the transforming growth factor (TGF)-ß family govern a wide range of mechanisms in brain development and in the adult, in particular neuronal/glial differentiation and survival, but also cell cycle regulation and neural stem cell maintenance. This clearly created some discrepancies in the field with some studies favouring neuronal differentiation/survival of progenitors and others favouring cell cycle exit and neural stem cell quiescence/maintenance. Here, we provide a unifying hypothesis claiming that through its regulation of neural progenitor cell (NPC) proliferation, TGF-ß signalling might be responsible for (i) maintaining stem cells in a quiescent stage, and (ii) promoting survival of newly generated neurons and their functional differentiation. Therefore, we performed a detailed histological analysis of TGF-ß1 signalling in the hippocampal neural stem cell niche of a transgenic mouse that was previously generated to express TGF-ß1 under a tetracycline regulatable Ca-Calmodulin kinase promoter. We also analysed NPC proliferation, quiescence, neuronal survival and differentiation in relation to elevated levels of TGF-ß1 in vitro and in vivo conditions. Finally, we performed a gene expression profiling to identify the targets of TGF-ß1 signalling in adult NPCs. The results demonstrate that TGF-ß1 promotes stem cell quiescence on one side, but also neuronal survival on the other side. Thus, considering the elevated levels of TGF-ß1 in ageing and neurodegenerative diseases, TGF-ß1 signalling presents a molecular target for future interventions in such conditions.

Age-dependent and differential effects of Smad7ΔEx1 on neural progenitor cell proliferation and on neurogenesis.

We recently reported that young (3 to 4months old) mice lacking Exon 1 of the Smad7 gene (S7ΔEx1 mice) show enhanced proliferation of neural stem and progenitor cells (NPCs) in the hippocampal dentate gyrus (DG) and in the subventricular zone (SVZ) of the lateral ventricles. It remained unclear, however, whether this phenotype would persist along aging, the latter typically being associated with a profound decrease in neurogenesis. Analysis of NPCs' proliferation based on the cell cycle marker PCNA in 12month-old S7ΔEx1 mice revealed a reversal of the phenotype. Hence, in contrast to their younger counterparts, 12month-old S7ΔEx1 mice had a reduced number of proliferating cells, compared to wildtype (WT) mice. At the same time, the survival of newly generated cells was enhanced in the aged transgenic animals. 12month-old S7ΔEx1 mice further displayed a reduced level of neurogenesis based on the numbers of cells expressing doublecortin (DCX), a marker for newborn neurons. The reduced neurogenesis in aged S7ΔEx1 mice was not due to a stem cell depletion, which might have occurred as a consequence of hyperproliferation in the young mice, since the number of Nestin and Sox2 positive cells was similar in WT and S7ΔEx1 mice. Instead, Nestin positive cells in the DG as well as primary neurosphere cultures derived from 12month-old S7ΔEx1 mice had a reduced capability to proliferate. However, after passaging, when released from their age- and niche-associated proliferative block, neurospheres from aged S7ΔEx1 mice regained the hyperproliferative property. Further, pSmad2 antibody staining intensity was elevated in the DG and SVZ of 12-month old transgenic compared to WT mice, indicating increased intracellular TGF-beta signaling in the aged S7ΔEx1 mice. In summary, this points toward differential effects of S7ΔEx1 on neurogenesis: (i) a hyperproliferation in young animals caused by a cell autonomous mechanism, and (ii) a TGF-beta dependent modulation of neurogenesis in aged S7ΔEx1 animals that abrogates the cell-intrinsic hyperproliferative properties and results in reduced proliferation, increased stem cell quiescence, and enhanced survival of newly generated cells.

Neurogenesis and neuronal regeneration in status epilepticus.

Neurogenesis in the adult central nervous system has been well documented in several mammals including humans. By now, a plethora of data has been generated with the aim of understanding the molecular and cellular events governing neurogenesis. This growing comprehension will provide the basis for modulation of neurogenesis for therapeutic purposes, in particular in neurodegenerative diseases. Herein, we review the current knowledge on neurogenesis, in particular in the frame of epilepsy, since seizures have massive effects on neurogenesis. Conversely, some studies have suggested that aberrant neurogenesis might contribute to the development or manifestation of epilepsy and, moreover, chronic inhibition of neurogenesis in epilepsy might contribute to comorbidities of epilepsy such as cognitive deficits. Therefore, a better understanding of neurogenesis in the context of epilepsy is still required for future therapeutic purposes.

The zebrafish myotome contains tonic muscle fibers: morphological characterization and time course of formation.

It is long known that the skeletal muscle of teleost fish contains muscle fibers which are in all probability of a tonic type according to morphological criteria. However, the evidence for the existence of teleost tonic fibers is still confined to a very small number of species, and knowledge concerning their ontogeny and possible functions is even more restricted. A remarkable deficit in this context is that it is not even exactly known whether the zebrafish, which is widely used to study vertebrate developmental biology, has such fibers, or how they arise. The present study demonstrates the existence of tonic fibers in the zebrafish myotome. They are identical with a fiber population previously termed "red muscle rim" fibers. A combined histochemical, immunocytochemical, and ultrastructural approach is used to characterize the morphology and development of these fibers. This study provides a basis for using the zebrafish model system in the future research on the developmental regulation and the functions of tonic fibers.

Inhibition of leukotriene receptors boosts neural progenitor proliferation.

Neural stem and progenitor cells serve as a reservoir for new neurons in the adult brain throughout lifetime. One of the critical steps determining the net production of new neurons is neural progenitor proliferation, which needs to be tightly controlled. Since inflammation has detrimental effects on neurogenesis and the 5-lipoxygenase/leukotriene pathway is involved in inflammatory processes, we investigated the effects of leukotrienes and montelukast, a small molecule inhibitor of the leukotriene receptors CysLT(1)R and GPR17, on neural stem and progenitor cell proliferation. We demonstrate expression of the leukotriene receptor GPR17 by neural progenitors and by neural stem cells. Stimulation with excess amounts of leukotrienes did not affect progenitor proliferation, whereas blockade of GPR17 with montelukast strongly elevated neural stem and progenitor proliferation, while maintaining their differentiation fate and potential. This effect was associated with increased ERK1/2 phosphorylation suggesting an involvement of the EGF signaling cascade. Based on our results, montelukast and the inhibition of the 5-LOX pathway might be potent candidates for future therapies employing neurogenesis to promote structural and functional improvement in neurodegeneration, neuropsychiatric disease and ageing.

The dark side of BrdU in neural stem cell biology: detrimental effects on cell cycle, differentiation and survival.

5-Bromo-2'-deoxyuridin (BrdU) is frequently used in anaylsis of neural stem cell biology, in particular to label and to fate-map dividing cells. However, up to now, only a few studies have addressed the question as to whether BrdU labeling per se affects the cells to be investigated. Here, we focused on the potential impact of BrdU on neurosphere cultures derived from the adult rat brain and on proliferation of progenitors in vivo. In vitro, neurospheres were pulsed for 48 h with BrdU, and cell proliferation, cell cycle, differentiation, survival and adhesion properties were subsequently analyzed. BrdU inhibited the expansion of neural progenitors as assessed by MTS assay and increased the fraction of cells in the G0/G1-phase of the cell cycle. Moreover, BrdU increased cell death and dose-dependently induced adherence of NPCs. Cell adherence was accompanied by a reduced amount of active matrix-metalloproteinase-2 (MMP-2). Furthermore, BrdU repressed neuronal and oligodendroglial differentiation, whereas astroglial fate was not affected. In contrast to the in vitro situation, BrdU apparently did not influence endogenous proliferation of NPCs or neurogenesis in concentrations that are typically used for labeling of neural progenitors in vivo. Our results reveal so far uncharacterized effects of BrdU on adult NPCs. We conclude that, because of its ubiquitous use in stem cell biology, any potential effect of BrdU of NPCs has to be scrutinized prior to interpretation of data.

Temperature-dependent modification of muscle precursor cell behaviour is an underlying reason for lasting effects on muscle cellularity and body growth of teleost fish.

Temperature is an important factor influencing teleost muscle growth, including a lasting ('imprinted') influence of embryonic thermal experience throughout all further life. However, little is known about the cellular processes behind this phenomenon. The study reported here used digital morphometry and immunolabelling for Pax7, myogenin and H3P to quantitatively examine the effects of thermal history on muscle precursor cell (MPC) behaviour and muscle growth in pearlfish (Rutilus meidingeri) until the adult stage. Fish were reared at three different temperatures (8.5, 13 and 16°C) until hatching and subsequently kept under the same (ambient) thermal conditions. Cellularity data were combined with a quantitative analysis of Pax7+ MPCs including those that were mitotically active (Pax7+/H3P+) or had entered differentiation (Pax7+/myogenin+). The results demonstrate that at hatching, body lengths, fast and slow muscle cross-sectional areas and fast fibre numbers are lower in fish reared at 8.5 and 13°C than at 16°C. During the larval period, this situation changes in the 13°C-fish, so that these fish are finally the largest. The observed effects can be related to divergent cellular mechanisms at the MPC level that are initiated in the embryo during the imprinting period. Embryos of 16°C-fish have reduced MPC proliferation but increased differentiation, and thus give rise to larger hatchlings. However, their limited MPC reserves finally lead to smaller adults. By contrast, embryos of 13°C-fish and, to a lesser extent, 8.5°-fish, show enhanced MPC proliferation but reduced differentiation, thus leading to smaller hatchlings but allowing for a larger MPC pool that can be used for enhanced post-hatching growth, finally resulting in larger adults.