Autocrine VEGF drives neural stem cell proximity to the adult hippocampus vascular niche.

The vasculature is a key component of adult brain neural stem cell (NSC) niches. In the adult mammalian hippocampus, NSCs reside in close contact with a dense capillary network. How this niche is maintained is unclear. We recently found that adult hippocampal NSCs express VEGF, a soluble factor with chemoattractive properties for vascular endothelia. Here, we show that global and NSC-specific VEGF loss led to dissociation of NSCs and their intermediate progenitor daughter cells from local vasculature. Surprisingly, though, we found no changes in local vascular density. Instead, we found that NSC-derived VEGF supports maintenance of gene expression programs in NSCs and their progeny related to cell migration and adhesion. In vitro assays revealed that blockade of VEGF receptor 2 impaired NSC motility and adhesion. Our findings suggest that NSCs maintain their own proximity to vasculature via self-stimulated VEGF signaling that supports their motility towards and/or adhesion to local blood vessels.

Excitatory amino acid transporter 1 supports adult hippocampal neural stem cell self-renewal.

Within the adult mammalian dentate gyrus (DG) of the hippocampus, glutamate stimulates neural stem cell (NSC) self-renewing proliferation, providing a link between adult neurogenesis and local circuit activity. Here, we show that glutamate-induced self-renewal of adult DG NSCs requires glutamate transport via excitatory amino acid transporter 1 (EAAT1) to stimulate lipogenesis. Loss of EAAT1 prevented glutamate-induced self-renewing proliferation of NSCs in vitro and in vivo, with little role evident for canonical glutamate receptors. Transcriptomics and further pathway manipulation revealed that glutamate simulation of NSCs relied on EAAT1 transport-stimulated lipogenesis. Our findings demonstrate a critical, direct role for EAAT1 in stimulating NSCs to support neurogenesis in adulthood, thereby providing insights into a non-canonical mechanism by which NSCs sense and respond to their niche.

Neural stem and progenitor cells support and protect adult hippocampal function via vascular endothelial growth factor secretion.

Adult neural stem and progenitor cells (NSPCs) reside in the dentate gyrus (DG) of the hippocampus throughout the lifespan of most mammalian species. In addition to generating new neurons, NSPCs may alter their niche via secretion of growth factors and cytokines. We recently showed that adult DG NSPCs secrete vascular endothelial growth factor (VEGF), which is critical for maintaining adult neurogenesis. Here, we asked whether NSPC-derived VEGF alters hippocampal function independent of adult neurogenesis. We found that loss of NSPC-derived VEGF acutely impaired hippocampal memory, caused neuronal hyperexcitability and exacerbated excitotoxic injury. We also found that NSPCs generate substantial proportions of total DG VEGF and VEGF disperses broadly throughout the DG, both of which help explain how this anatomically-restricted cell population could modulate function broadly. These findings suggest that NSPCs actively support and protect DG function via secreted VEGF, thereby providing a non-neurogenic functional dimension to endogenous NSPCs.

Estimation of the density of neural, glial, and endothelial lineage cells in the adult mouse dentate gyrus.

The dentate gyrus subregion of the mammalian hippocampus is an adult neural stem cell niche and site of lifelong neurogenesis. Hypotheses regarding the role of adult-born neuron synaptic integration in hippocampal circuit function are framed by robust estimations of adult-born versus pre/perinatally-born neuron number. In contrast, the non-neurogenic functions of adult neural stem cells and their immediate progeny, such as secretion of bioactive growth factors and expression of extracellular matrix-modifying proteins, lack similar framing due to few estimates of their number versus other prominent secretory cells. Here, we apply immunohistochemical methods to estimate cell density of neural stem/progenitor cells versus other major classes of glial and endothelial cell types that are potentially secretory in the dentate gyrus of adult mice. Of the cell types quantified, we found that GFAP+SOX2+ stellate astrocytes were the most numerous, followed by CD31+ endothelia, GFAP-SOX2+ intermediate progenitors, Olig2+ oligodendrocytes, Iba1+ microglia, and GFAP+SOX2+ radial glia-like neural stem cells. We did not observe any significant sex differences in density of any cell population. Notably, neural stem/progenitor cells were present at a similar density as several cell types known to have potent functional roles via their secretome. These findings may be useful for refining hypotheses regarding the contributions of these cell types to regulating hippocampal function and their potential therapeutic uses. All experimental protocols were approved by the Ohio State University Institutional Animal Care and Use Committee (protocol# 2016A00000068) on July 14, 2016.

Editorial: Stem cell secretome.

Stem cells have the potential to advance therapy for many neurological diseases that are currently refractive to treatment. They are also key cellular players in homeostasis within several adult brain regions that host endogenous populations of neural stem cells. Investigations of the functions of stem cells in the adult CNS have historically approached these cells as sources of differentiated progeny, whether it be new neurons or new glial cells. Yet, as both basic research and pre-clinical efforts centered on stem cells in the brain push forward, it has become evident that this initial framework is incomplete. Emerging evidence indicates that stem and progenitor cells from a variety of tissues can regulate their microenvironment through production of secreted factors. This special issue highlights work investigating the role of the neural and non-neural stem cell secretome in regulating CNS function. These studies represent efforts both to more fully delineate the suite of factors secreted by stem cells and to evaluate its impact on CNS health and disease. Together, they demonstrate a broad potential for stem cell function through secreted proteins that urges continued basic and translational research in the years to come.

Defining the adult hippocampal neural stem cell secretome: In vivo versus in vitro transcriptomic differences and their correlation to secreted protein levels.

Adult hippocampal neural stem and progenitor cells (NSPCs) secrete a variety of proteins that affect tissue function. Though several individual NSPC-derived proteins have been shown to impact key cellular processes, a broad characterization is lacking. Secretome profiling of low abundance stem cell populations is typically achieved via proteomic characterization of in vitro, isolated cells. Here, we identified hundreds of secreted proteins in conditioned media from in vitro adult mouse hippocampal NSPCs using an antibody array and mass spectrometry. Comparison of protein abundance between antibody array and mass spectrometry plus quantification of several key secreted proteins by ELISA revealed notable disconnect between methods in what proteins were identified as being high versus low abundance, suggesting that data from antibody arrays in particular should be approached with caution. We next assessed the NSPC secretome on a transcriptional level with single cell and bulk RNA sequencing (RNAseq) of cultured NSPCs. Comparison of RNAseq transcript levels of highly secreted proteins revealed that quantification of gene expression did not necessarily predict relative protein abundance. Interestingly, comparing our in vitro NSPC gene expression data with similar data from freshly isolated, in vivo hippocampal NSPCs revealed strong correlations in global gene expression between in vitro and in vivo NSPCs. Understanding the components and functions of the NSPC secretome is essential to understanding how these cells may modulate the hippocampal neurogenic niche. Cumulatively, our data emphasize the importance of using proteomics in conjunction with transcriptomics and highlights the need for better methods of unbiased secretome profiling.

Threats from the past: Barbados green monkeys (Chlorocebus sabaeus) fear leopards after centuries of isolation.

Ability to recognize and differentiate between predators and non-predators is a crucial component of successful anti-predator behavior. While there is evidence that both genetic and experiential mechanisms mediate anti-predator behaviors in various animal species, it is unknown to what extent each of these two mechanisms are utilized by the green monkey (Chlorocebus sabaeus). Green monkeys on the West Indies island of Barbados offer a unique opportunity to investigate the underpinnings of anti-predator behaviors in a species that has been isolated from ancestral predators for over 350 years. In the first experiment, monkeys in two free-ranging troops were presented with photographs of an ancestral predator (leopard, Panthera pardus) and a non-predator (African Buffalo, Syncerus caffer). Relative to non-predator stimuli, images of a leopard elicited less approach, more alarm calls, and more escape responses. Subsequent experiments were conducted to determine whether the monkeys were responding to a leopard-specific feature (spotted fur) or a general predator feature (forward facing eyes). The monkeys showed similar approach to images of an unfamiliar non-predator regardless of whether the image had forward facing predator eyes or side facing non-predator eyes. However, once near the images, the monkeys were less likely to reach for peanuts near the predator eyes than the non-predator eyes. The monkeys avoided an image of spotted leopard fur but approached the same image of fur when the dark spots had been removed. Taken together, the results suggest that green monkey anti-predator behavior is at least partially mediated by genetic factors.