German Center for Neurodegenerative Diseases (DZNE) Dresden

Several concepts, which in the aggregate get might be used to account for "resilience" against age- and disease-related changes, have been the subject of much research. These include brain reserve, cognitive reserve, and brain maintenance. However, different investigators have use these terms in different ways, and there has never been an attempt to arrive at consensus on the definition of these concepts. Furthermore, there has been confusion regarding the measurement of these constructs and the appropriate ways to apply them to research. Therefore the reserve, resilience, and protective factors professional interest area, established under the auspices of the Alzheimer's Association, established a whitepaper workgroup to develop consensus definitions for cognitive reserve, brain reserve, and brain maintenance. The workgroup also evaluated measures that have been used to implement these concepts in research settings and developed guidelines for research that explores or utilizes these concepts. The workgroup hopes that this whitepaper will form a reference point for researchers in this area and facilitate research by supplying a common language.

Of the neurogenic zones in the adult brain, adult hippocampal neurogenesis attracts the most attention, because it is involved in higher cognitive function, most notably memory processes, and certain affective behaviors. Adult hippocampal neurogenesis is also found in humans at a considerable level and appears to contribute significantly to hippocampal plasticity across the life span, because it is regulated by activity. Adult hippocampal neurogenesis generates new excitatory granule cells in the dentate gyrus, whose axons form the mossy fiber tract that links the dentate gyrus to CA3. It originates from a population of radial glia-like precursor cells (type 1 cells) that have astrocytic properties, express markers of neural stem cells and divide rarely. They give rise to intermediate progenitor cells with first glial (type 2a) and then neuronal (type 2b) phenotype. Through a migratory neuroblast-like stage (type 3), the newborn, lineage-committed cells exit the cell cycle and enter a maturation stage, during which they extend their dendrites into a the molecular layer and their axon to CA3. They go through a period of several weeks, during which they show increased synaptic plasticity, before finally becoming indistinguishable from the older granule cells.

The plasticity of pluripotent stem cells provides new possibilities for studying development, degeneration, and regeneration. Protocols for the differentiation of retinal organoids from embryonic stem cells have been developed, which either recapitulate complete eyecup morphogenesis or maximize photoreceptor genesis. Here, we have developed a protocol for the efficient generation of large, 3D-stratified retinal organoids that does not require evagination of optic-vesicle-like structures, which so far limited the organoid yield. Analysis of gene expression in individual organoids, cell birthdating, and interorganoid variation indicate efficient, reproducible, and temporally regulated retinogenesis. Comparative analysis of a transgenic reporter for PAX6, a master regulator of retinogenesis, shows expression in similar cell types in mouse in vivo, and in mouse and human retinal organoids. Early or late Notch signaling inhibition forces cell differentiation, generating organoids enriched with cone or rod photoreceptors, respectively, demonstrating the power of our improved organoid system for future research in stem cell biology and regenerative medicine.

Voluntary wheel running has long been known to induce precursor cell proliferation in adult hippocampal neurogenesis in rodents. However, mechanisms that couple activity with the promitotic effect are not yet fully understood. Using tryptophan hydroxylase (TPH) 2 deficient (Tph2-deficient) mice that lack brain serotonin, we explored the relationship between serotonin signaling and exercise-induced neurogenesis. Surprisingly, Tph2-deficient mice exhibit normal baseline hippocampal neurogenesis but impaired activity-induced proliferation. Our data demonstrate that the proproliferative effect of running requires the release of central serotonin in young-adult and aged mice. Lack of brain serotonin further results in alterations at the stage of Sox2-positive precursor cells, suggesting physiological adaptations to changes in serotonin supply to maintain homeostasis in the neurogenic niche. We conclude that serotonin plays a direct and acute regulatory role in activity-dependent hippocampal neurogenesis. The understanding of exercise-induced neurogenesis might offer preventive but also therapeutic opportunities in depression and age-related cognitive decline.

The Morris water maze represents the de-facto standard for testing hippocampal function in laboratory rodents. In the field of adult hippocampal neurogenesis, however, using this paradigm to assess the functional relevance of the new neurons yielded surprisingly inconsistent results. While some authors found aspects of water maze performance to be linked to adult neurogenesis, others obtained different results or could not demonstrate any effect of manipulating adult neurogenesis. In this review we discuss evidence that the large diversity of protocols and setups used is an important aspect in interpreting the differences in the results that have been obtained. Even simple parameters such as pool size, number, and configuration of visual landmarks, or number of trials can become highly relevant for getting the new neurons involved at all. Sets of parameters are often chosen with implicit or explicit concepts in mind and these might lead to different views on the function of adult-generated neurons. We propose that the classical parameters usually used to measure spatial learning performance in the water maze might not be particularly well-suited to sensitively and specifically detect the supposedly highly specific functional changes elicited by the experimental modulation of adult hippocampal neurogenesis. As adult neurogenesis is supposed to affect specific aspects of information processing only in the hippocampus, any claim for a functional relevance of the new neurons has to be based on hippocampus-specific parameters. We also placed a special emphasis on the fact that the dentate gyrus (DG) facilitates the differentiation between contexts as opposed to just differentiating places. In conclusion, while the Morris water maze has proven to be one of the most effective testing paradigms to assess hippocampus-dependent spatial learning, new and more specific questions ask for new parameters. Therefore, the full potential of the water maze task remains to be tapped.

We here show that living in a stimulus-rich environment (ENR) improves water maze learning with respect to specific key indicators that in previous loss-of-function experiments have been shown to rely on adult hippocampal neurogenesis. Analyzing the strategies employed by mice to locate the hidden platform in the water maze revealed that ENR facilitated task acquisition by increasing the probability to use effective search strategies. ENR also enhanced the animals' behavioral flexibility, when the escape platform was moved to a new location. Treatment with temozolomide, which is known to reduce adult neurogenesis, abolished the effects of ENR on both acquisition and flexibility, while leaving other aspects of water maze learning untouched. These characteristic effects and interdependencies were not seen in parallel experiments with voluntary wheel running (RUN), a second pro-neurogenic behavioral stimulus. Since the histological assessment of adult neurogenesis is by necessity an end-point measure, the levels of neurogenesis over the course of the experiment can only be inferred and the present study focused on behavioral parameters as analytical endpoints. Although the correlation of physical activity with precursor cell proliferation and of learning and the survival of new neurons is well established, how the specific functional effects described here relate to dynamic changes in the stem cell niche remains to be addressed. Nevertheless, our findings support the hypothesis that adult neurogenesis is a critical mechanism underlying the beneficial effects of leading an active live, rich in experiences.

INTRODUCTION: White matter hyperintensities (WMH) are often described in Alzheimer's disease (AD), but their topography and specific relationships with cognition remain unclear. METHODS: Regional WMH were estimated in 54 cognitively impaired amyloid beta-positive AD (Aβpos-AD), compared to 40 cognitively unimpaired amyloid beta-negative older controls (Aβneg-controls) matched for vascular risk factors. The cross-sectional association between regional WMH volume and cognition was assessed within each group, controlling for cerebral amyloid burden, global cortical atrophy, and hippocampal atrophy. RESULTS: WMH volume was larger in Aβpos-AD compared to Aβneg-controls in all regions, with the greatest changes in the splenium of the corpus callosum (S-CC). In Aβpos-AD patients, larger total and regional WMH volume, especially in the S-CC, was strongly associated with decreased cognition. DISCUSSION: WMH specifically contribute to lower cognition in AD, independently from amyloid deposition and atrophy. This study emphasizes the clinical relevance of WMH in AD, especially posterior WMH, and most notably S-CC WMH.

Although the neurogenesis-enhancing effects of exercise have been extensively studied, the molecular mechanisms underlying this response remain unclear. Here, we propose that this is mediated by the exercise-induced systemic release of the antioxidant selenium transport protein, selenoprotein P (SEPP1). Using knockout mouse models, we confirmed that SEPP1 and its receptor low-density lipoprotein receptor-related protein 8 (LRP8) are required for the exercise-induced increase in adult hippocampal neurogenesis. In vivo selenium infusion increased hippocampal neural precursor cell (NPC) proliferation and adult neurogenesis. Mimicking the effect of exercise through dietary selenium supplementation restored neurogenesis and reversed the cognitive decline associated with aging and hippocampal injury, suggesting potential therapeutic relevance. These results provide a molecular mechanism linking exercise-induced changes in the systemic environment to the activation of quiescent hippocampal NPCs and their subsequent recruitment into the neurogenic trajectory.

OBJECTIVE: Weight loss has been identified as a negative prognostic factor in amyotrophic lateral sclerosis, but there is no evidence regarding whether a high-caloric diet increases survival. Therefore, we sought to evaluate the efficacy of a high-caloric fatty diet (HCFD) for increasing survival. METHODS: A 1:1 randomized, placebo-controlled, parallel-group, double-blinded trial (LIPCAL-ALS study) was conducted between February 2015 and September 2018. Patients were followed up at 3, 6, 9, 12, 15, and 18 months after randomization. The study was performed at 12 sites of the clinical and scientific network of German motor neuron disease centers (ALS/MND-NET). Eligible patients were randomly assigned (1:1) to receive either HCFD (405kcal/day, 100% fat) or placebo in addition to riluzole (100mg/day). The primary endpoint was survival time, defined as time to death or time to study cutoff date. RESULTS: Two hundred one patients (80 female, 121 male, age = 62.4 ± 10.8 years) were included. The confirmatory analysis of the primary outcome survival showed a survival probability of 0.39 (95% confidence interval [CI] = 0.27-0.51) in the placebo group and 0.37 (95% CI = 0.25-0.49) in the HCFD group, both after 28 months (point in time of the last event). The hazard ratio was 0.97, 1-sided 97.5% CI = -∞ to 1.44, p = 0.44. INTERPRETATION: The results provide no evidence for a life-prolonging effect of HCFD for the whole amyotrophic lateral sclerosis population. However, post hoc analysis revealed a significant survival benefit for the subgroup of fast-progressing patients. ANN NEUROL 2020;87:206-216.

When does adult hippocampal neurogenesis begin? We describe the development of the neurogenic niche in the subgranular zone (SGZ) of the hippocampal dentate gyrus. We did so from the perspective of the situation in the adult. Ontogeny of the dentate gyrus is complex and results in an ectopic neurogenic niche that lifelong generates new granule cells. Neurogenesis during the fetal and early postnatal periods builds the dentate gyrus and gives way to activity-dependent "adult" neurogenesis. We used markers most relevant to adult neurogenesis research to describe this transition: Nestin, Sox2, BLBP, GFAP, Tbr2, Doublecortin (DCX), NeuroD1 and Prox1. We found that massive changes and a local condensation of proliferating precursor cells occurs between postnatal day 7 (P7), near the peak in proliferation, and P14. Before and around P7, the spatial distribution of cells and the co-localization of markers were distinct from the situation in the adult. Unlike the adult SGZ, the marker pair Nestin/Sox2 and the radial glial marker BLBP were not overlapping during embryonic development, presumably indicating different types of radial glia-like cells. Before P7 GFAP-positive cells in the hilus lacked the radial orientation that is characteristic of the adult type-1 cells. DCX, which is concentrated in type-2b and type-3 progenitor cells and early postmitotic neurons in the adult, showed diffuse expression before P7. Intermediate progenitor cell marker Tbr2 became restricted to the SGZ but was found in the granule cell layer (GCL) and hilus before. Lineage markers NeuroD1 and Prox1 confirmed this pattern. We conclude that the neurogenic niche of adult neurogenesis is in place well before true adulthood. This might indicate that consistent with the hypothesized function of adult neurogenesis in activity-dependent plasticity, the early transition from postnatal neurogenesis to adult neurogenesis coincides with the time, when the young mice start to become active themselves.

Brain activity and connectivity alter drastically during epileptic seizures. The brain networks shift from a balanced resting state to a hyperactive and hypersynchronous state. It is, however, less clear which mechanisms underlie the state transitions. By studying neural and glial activity in zebrafish models of epileptic seizures, we observe striking differences between these networks. During the preictal period, neurons display a small increase in synchronous activity only locally, while the gap-junction-coupled glial network was highly active and strongly synchronized across large distances. The transition from a preictal state to a generalized seizure leads to an abrupt increase in neural activity and connectivity, which is accompanied by a strong alteration in glia-neuron interactions and a massive increase in extracellular glutamate. Optogenetic activation of glia excites nearby neurons through the action of glutamate and gap junctions, emphasizing a potential role for glia-glia and glia-neuron connections in the generation of epileptic seizures.

A major cause for vision impairment and blindness in industrialized countries is the loss of the light-sensing retinal tissue in the eye. Photoreceptor damage is one of the main characteristics found in retinal degeneration diseases, such as Retinitis Pigmentosa or age-related macular degeneration. The lack of effective therapies to stop photoreceptor loss together with the absence of significant intrinsic regeneration in the human retina converts such degenerative diseases into permanent conditions that are currently irreversible. Cell replacement by means of photoreceptor transplantation has been proposed as a potential approach to tackle cell loss in the retina. Since the first attempt of photoreceptor transplantation in humans, about twenty years ago, several research groups have focused in the development and improvement of technologies necessary to bring cell transplantation for retinal degeneration diseases to reality. Progress in recent years in the generation of human tissue derived from pluripotent stem cells (PSCs) has significantly improved our tools to study human development and disease in the dish. Particularly the availability of 3D culture systems for the generation of PSC-derived organoids, including the human retina, has dramatically increased access to human material for basic and medical research. In this review, we focus on important milestones towards the generation of transplantable photoreceptor precursors from PSC-derived retinal organoids and discuss recent pre-clinical transplantation studies using organoid-derived photoreceptors in context to related in vivo work using primary photoreceptors as donor material. Additionally, we summarize remaining challenges for developing photoreceptor transplantation towards clinical application.

The neurosphere assay and the adherent monolayer culture system are valuable tools to determine the potential (proliferation or differentiation) of adult neural stem cells in vitro. These assays can be used to compare the precursor potential of cells isolated from genetically different or differentially treated animals to determine the effects of exogenous factors on neural precursor cell proliferation and differentiation and to generate neural precursor cell lines that can be assayed over continuous passages. The neurosphere assay is traditionally used for the post-hoc identification of stem cells, primarily due to the lack of definitive markers with which they can be isolated from primary tissue and has the major advantage of giving a quick estimate of precursor cell numbers in brain tissue derived from individual animals. Adherent monolayer cultures, in contrast, are not traditionally used to compare proliferation between individual animals, as each culture is generally initiated from the combined tissue of between 5-8 animals. However, they have the major advantage that, unlike neurospheres, they consist of a mostly homogeneous population of precursor cells and are useful for following the differentiation process in single cells. Here, we describe, in detail, the generation of neurosphere cultures and, for the first time, adherent cultures from individual animals. This has many important implications including paired analysis of proliferation and/or differentiation potential in both the subventricular zone (SVZ) and dentate gyrus (DG) of treated or genetically different mouse lines, as well as a significant reduction in animal usage.

The neural stem cell (NSC) reservoir can be harnessed for stem cell-based regenerative therapies. Zebrafish remarkably regenerate their brain by inducing NSC plasticity in a Amyloid-β-42 (Aβ42)-induced experimental Alzheimer's disease (AD) model. Interleukin-4 (IL-4) is also critical for AD-induced NSC proliferation. However, the mechanisms of this response have remained unknown. Using single-cell transcriptomics in the adult zebrafish brain, we identify distinct subtypes of NSCs and neurons and differentially regulated pathways and their gene ontologies and investigate how cell-cell communication is altered through ligand-receptor pairs in AD conditions. Our results propose the existence of heterogeneous and spatially organized stem cell populations that react distinctly to amyloid toxicity. This resource article provides an extensive database for the molecular basis of NSC plasticity in the AD model of the adult zebrafish brain. Further analyses of stem cell heterogeneity and neuro-regenerative ability at single-cell resolution could yield drug targets for mobilizing NSCs for endogenous neuro-regeneration in humans.

Cellular redox states regulate the balance between stem cell maintenance and activation. Increased levels of intracellular reactive oxygen species (ROS) are linked to proliferation and lineage specification. In contrast to this general principle, we here show that in the hippocampus of adult mice, quiescent neural precursor cells (NPCs) maintain the highest ROS levels (hiROS). Classifying NPCs on the basis of cellular ROS content identified distinct functional states. Shifts in ROS content primed cells for a subsequent state transition, with lower ROS content marking proliferative activity and differentiation. Physical activity, a physiological activator of adult hippocampal neurogenesis, recruited hiROS NPCs into proliferation via a transient Nox2-dependent ROS surge. In the absence of Nox2, baseline neurogenesis was unaffected, but the activity-induced increase in proliferation disappeared. These results provide a metabolic classification of NPC functional states and describe a mechanism linking the modulation of cellular ROS by behavioral cues to the activation of adult NPCs.

Physical activity is a strong positive physiological modulator of adult neurogenesis in the hippocampal dentate gyrus. Although the underlying regulatory mechanisms are still unknown, systemic processes must be involved. Here we show that platelets are activated after acute periods of running, and that activated platelets promote neurogenesis, an effect that is likely mediated by platelet factor 4. Ex vivo, the beneficial effects of activated platelets and platelet factor 4 on neural precursor cells were dentate gyrus specific and not observed in the subventricular zone. Moreover, the depletion of circulating platelets in mice abolished the running-induced increase in precursor cell proliferation in the dentate gyrus following exercise. These findings demonstrate that platelets and their released factors can modulate adult neural precursor cells under physiological conditions and provide an intriguing link between running-induced platelet activation and the modulation of neurogenesis after exercise.

BACKGROUND: The concept of reserve was established to account for the observation that a given degree of neurodegenerative pathology may result in varying degrees of symptoms in different individuals. There is a large amount of evidence on epidemiological risk and protective factors for neurodegenerative diseases and dementia, yet the biological mechanisms that underpin the protective effects of certain lifestyle and physiological variables remain poorly understood, limiting the development of more effective preventive and treatment strategies. Additionally, different definitions and concepts of reserve exist, which hampers the coordination of research and comparison of results across studies. DISCUSSION: This paper represents the consensus of a multidisciplinary group of experts from different areas of research related to reserve, including clinical, epidemiological and basic sciences. The consensus was developed during meetings of the working groups of the first International Conference on Cognitive Reserve in the Dementias (24-25 November 2017, Munich, Germany) and the Alzheimer's Association Reserve and Resilience Professional Interest Area (25 July 2018, Chicago, USA). The main objective of the present paper is to develop a translational perspective on putative mechanisms underlying reserve against neurodegenerative disease, combining evidence from epidemiological and clinical studies with knowledge from animal and basic research. The potential brain functional and structural basis of reserve in Alzheimer's disease and other brain disorders are discussed, as well as relevant lifestyle and genetic factors assessed in both humans and animal models. CONCLUSION: There is an urgent need to advance our concept of reserve from a hypothetical model to a more concrete approach that can be used to improve the development of effective interventions aimed at preventing dementia. Our group recommends agreement on a common dictionary of terms referring to different aspects of reserve, the improvement of opportunities for data sharing across individual cohorts, harmonising research approaches across laboratories and groups to reduce heterogeneity associated with human data, global coordination of clinical trials to more effectively explore whether reducing epidemiological risk factors leads to a reduced burden of neurodegenerative diseases in the population, and an increase in our understanding of the appropriateness of animal models for reserve research.

Age and activity might be considered the two antagonistic key regulators of adult neurogenesis. Adult neurogenesis decreases with age but remains present, albeit at a very low level, even in the oldest individuals. Activity, be it physical or cognitive, increases adult neurogenesis and thereby seems to counteract age effects. It is, thus, proposed that activity-dependent regulation of adult neurogenesis might contribute to some sort of "neural reserve," the brain's ability to compensate functional loss associated with aging or neurodegeneration. Activity can have nonspecific and specific effects on adult neurogenesis. Mechanistically, nonspecific stimuli that largely affect precursor cell stages might be related by the local microenvironment, whereas more specific, survival-promoting effects take place at later stages of neuronal development and require the synaptic integration of the new cell and its particular synaptic plasticity.

We need to effectively combine the knowledge from surging literature with complex datasets to propose mechanistic models of SARS-CoV-2 infection, improving data interpretation and predicting key targets of intervention. Here, we describe a large-scale community effort to build an open access, interoperable and computable repository of COVID-19 molecular mechanisms. The COVID-19 Disease Map (C19DMap) is a graphical, interactive representation of disease-relevant molecular mechanisms linking many knowledge sources. Notably, it is a computational resource for graph-based analyses and disease modelling. To this end, we established a framework of tools, platforms and guidelines necessary for a multifaceted community of biocurators, domain experts, bioinformaticians and computational biologists. The diagrams of the C19DMap, curated from the literature, are integrated with relevant interaction and text mining databases. We demonstrate the application of network analysis and modelling approaches by concrete examples to highlight new testable hypotheses. This framework helps to find signatures of SARS-CoV-2 predisposition, treatment response or prioritisation of drug candidates. Such an approach may help deal with new waves of COVID-19 or similar pandemics in the long-term perspective.

When adult neurogenesis was discovered in the mammalian brain it was often considered an atavism and, even today, many people are convinced that there has been a "phylogenetic reduction" away from lifelong neurogenesis, favoring stability for complex brains. Adult neurogenesis is found throughout the animal kingdom but varies to a large extent. Mammals might have fewer neurogenic zones than, for example, fish, but within their remaining neurogenic zones, the new neurons are highly functional. Especially, humans have very substantial quantities of neurogenesis in their hippocampus. At least for the mammalian dentate gyrus, one can thus argue that there has been evolution toward neurogenesis-based plasticity rather than away from it.