βB2-crystallin (gene symbol: Crybb2/CRYBB2) was first described as a structural protein of the ocular lens. This gene, however, is also expressed in several regions of the mammalian brain, although its function in this organ remains entirely unknown. To unravel some aspects of its function in the brain, we combined behavioral, neuroanatomical, and physiological analyses in a novel Crybb2 mouse mutant, O377. Behavioral tests with male O377 mutants revealed altered sensorimotor gating, suggesting modified neuronal functions. Since these mouse mutants also displayed reduced hippocampal size, we concentrated further investigations on the hippocampus. Free intracellular Ca(2+) levels were increased and apoptosis was enhanced in the hippocampus of O377 mutants. Moreover, the expression of the gene encoding calpain 3 (gene symbol Capn3) was elevated and the expression of genes coding for the NMDA receptor subunits was downregulated. Additionally, the number of parvalbumin-positive interneurons was decreased in the hippocampus but not in the cortex of the mutants. High-speed voltage-sensitive dye imaging demonstrated an increased translation of input-to-output neuronal activity in the dentate gyrus of this Crybb2 mutant. These results point to an important function of βB2-crystallin in the hippocampal network. They indicate pleiotropic effects of mutations in the Crybb2 gene, which previously had been considered to be specific to the ocular lens. Moreover, our results are the first to demonstrate that βB2-crystallin has a role in hippocampal function and behavioral phenotypes. This model can now be further explored by future experiments.

The role played by adult neurogenesis in anxiety is not clear. A recent study revealed a surprising positive correlation between increased anxiety and elevated neurogenesis following chronic voluntary wheel running and multiple behavioural testing in mice, suggesting that adult hippocampal neurogenesis is involved in the genesis of anxiety. To exclude the possible confounding effect of multiple testing that may have occurred in the aforementioned study, we assessed (1) the effects of mouse voluntary wheel running (14 vs. 28 days) on anxiety in just one behavioural test; the open field, and (2), using different markers, proliferation, differentiation, survival and maturation of newly born neurons in the dentate gyrus immediately afterwards. Effects of wheel running on anxiety-related behaviour were confirmed in a separate batch of animals tested in another test of anxiety, the light/dark box test.

Neurogenesis in the brain of adult mammals occurs throughout life in two locations: the subventricular zone of the lateral ventricle and the subgranular zone of the dentate gyrus in the hippocampus. RNA interference mechanisms have emerged as critical regulators of neuronal differentiation. However, to date, little is known about its function in adult neurogenesis.

Haploinsufficiency of the class I bHLH transcription factor TCF4 causes Pitt-Hopkins syndrome (PTHS), a severe neurodevelopmental disorder, while common variants in the TCF4 gene have been identified as susceptibility factors for schizophrenia. It remains largely unknown, which brain regions are dependent on TCF4 for their development and function.

Transcription factor 4 (TCF4) has been linked to human neurodevelopmental disorders such as intellectual disability, Pitt-Hopkins Syndrome (PTHS), autism, and schizophrenia. Recent work demonstrated that TCF4 participates in the control of a wide range of neurodevelopmental processes in mammalian nervous system development including neural precursor proliferation, timing of differentiation, migration, dendritogenesis and synapse formation. TCF4 is highly expressed in the adult hippocampal dentate gyrus - one of the few brain regions where neural stem / progenitor cells generate new functional neurons throughout life.

Mitochondria are key organelles in regulating the metabolic state of a cell. In the brain, mitochondrial oxidative metabolism is the prevailing mechanism for neurons to generate ATP. While it is firmly established that neuronal function is highly dependent on mitochondrial metabolism, it is less well-understood how astrocytes function rely on mitochondria. In this study, we investigate if astrocytes require a functional mitochondrial electron transport chain (ETC) and oxidative phosphorylation (oxPhos) under physiological and injury conditions. By immunohistochemistry we show that astrocytes expressed components of the ETC and oxPhos complexes in vivo. Genetic inhibition of mitochondrial transcription by conditional deletion of mitochondrial transcription factor A (Tfam) led to dysfunctional ETC and oxPhos activity, as indicated by aberrant mitochondrial swelling in astrocytes. Mitochondrial dysfunction did not impair survival of astrocytes, but caused a reactive gliosis in the cortex under physiological conditions. Photochemically initiated thrombosis induced ischemic stroke led to formation of hyperfused mitochondrial networks in reactive astrocytes of the perilesional area. Importantly, mitochondrial dysfunction significantly reduced the generation of new astrocytes and increased neuronal cell death in the perilesional area. These results indicate that astrocytes require a functional ETC and oxPhos machinery for proliferation and neuroprotection under injury conditions.

Neuronal production from neural stem cells persists during adulthood in the subgranular zone of the hippocampal dentate gyrus. Extracellular signals provided by the hippocampal microenvironment regulate the neuronal fate commitment of the stem cell progeny. To date, the identity of those signals and their crosstalk has been only partially resolved. Here we show that adult rat hippocampal neural stem and progenitor cells (AH-NSPCs) express receptors for bone morphogenetic proteins (BMPs) and that the BMP/P-Smad pathway is active in AH-NSPCs undergoing differentiation towards the neuronal lineage. In vitro, exposure to the BMP2 and BMP4 ligands is sufficient to increase neurogenesis from AH-NSPCs in a WNT dependent manner while decreasing oligodendrogenesis. Moreover, BMP2/4 and WNT3A, a key regulator of adult hippocampal neurogenesis, cooperate to further enhance neuronal production. Our data point to a mechanistic convergence of the BMP and WNT pathways at the level of the T-cell factor/lymphoid enhancer factor gene Lef1. Altogether, we provide evidence that BMP signalling is an important regulator for the neuronal fate specification of AH-NSPCs cultures and we show that it significantly cooperates with the previously described master regulator of adult hippocampal neurogenesis, the WNT signalling pathway.

The high-mobility-group domain containing SoxC transcription factors Sox4 and Sox11 are expressed and required in the vertebrate central nervous system in neuronal precursors and neuroblasts. To identify genes that are widely regulated by SoxC proteins during vertebrate neurogenesis we generated expression profiles from developing mouse brain and chicken neural tube with reduced SoxC expression and found the transcription factor prospero homeobox protein 1 (Prox1) strongly down-regulated under both conditions. This led us to hypothesize that Prox1 expression depends on SoxC proteins in the developing central nervous system of mouse and chicken. By combining luciferase reporter assays and over-expression in the chicken neural tube with in vivo and in vitro binding studies, we identify the Prox1 gene promoter and two upstream enhancers at -44 kb and -40 kb relative to the transcription start as regulatory regions that are bound and activated by SoxC proteins. This argues that Prox1 is a direct target gene of SoxC proteins during neurogenesis. Electroporations in the chicken neural tube furthermore show that Prox1 activates a subset of SoxC target genes, whereas it has no effects on others. We propose that the transcriptional control of Prox1 by SoxC proteins may ensure coupling of two types of transcription factors that are both required during early neurogenesis, but have at least in part distinct functions. Open Data: Materials are available on https://cos.io/our-services/open-science-badges/ https://osf.io/93n6m/.

Autophagy is a conserved catabolic pathway with emerging functions in mammalian neurodevelopment and human neurodevelopmental diseases. The mechanisms controlling autophagy in neuronal development are not fully understood. Here, we found that conditional deletion of the Forkhead Box O transcription factors FoxO1, FoxO3, and FoxO4 strongly impaired autophagic flux in developing neurons of the adult mouse hippocampus. Moreover, FoxO deficiency led to altered dendritic morphology, increased spine density, and aberrant spine positioning in adult-generated neurons. Strikingly, pharmacological induction of autophagy was sufficient to correct abnormal dendrite and spine development of FoxO-deficient neurons. Collectively, these findings reveal a novel link between FoxO transcription factors, autophagic flux, and maturation of developing neurons.

Precise regulation of cellular metabolism is hypothesized to constitute a vital component of the developmental sequence underlying the life-long generation of hippocampal neurons from quiescent neural stem cells (NSCs). The identity of stage-specific metabolic programs and their impact on adult neurogenesis are largely unknown. We show that the adult hippocampal neurogenic lineage is critically dependent on the mitochondrial electron transport chain and oxidative phosphorylation machinery at the stage of the fast proliferating intermediate progenitor cell. Perturbation of mitochondrial complex function by ablation of the mitochondrial transcription factor A (Tfam) reproduces multiple hallmarks of aging in hippocampal neurogenesis, whereas pharmacological enhancement of mitochondrial function ameliorates age-associated neurogenesis defects. Together with the finding of age-associated alterations in mitochondrial function and morphology in NSCs, these data link mitochondrial complex function to efficient lineage progression of adult NSCs and identify mitochondrial function as a potential target to ameliorate neurogenesis-defects in the aging hippocampus.

The transcription factor FOXG1 serves pleiotropic functions in brain development ranging from the regulation of precursor proliferation to the control of cortical circuit formation. Loss-of-function mutations and duplications of FOXG1 are associated with neurodevelopmental disorders in humans illustrating the importance of FOXG1 dosage for brain development. Aberrant FOXG1 dosage has been found to disrupt the balanced activity of glutamatergic and GABAergic neurons, but the underlying mechanisms are not fully understood. We report that FOXG1 is expressed in the main adult neurogenic niches in mice, i.e. the hippocampal dentate gyrus and the subependymal zone/olfactory bulb system, where neurogenesis of glutamatergic and GABAergic neurons persists into adulthood. These niches displayed differential vulnerability to increased FOXG1 dosage: high FOXG1 levels severely compromised survival and glutamatergic dentate granule neuron fate acquisition in the hippocampal neurogenic niche, but left neurogenesis of GABAergic neurons in the subependymal zone/olfactory bulb system unaffected. Comparative transcriptomic analyses revealed a significantly higher expression of the apoptosis-linked nuclear receptor Nr4a1 in FOXG1-overexpressing hippocampal neural precursors. Strikingly, pharmacological interference with NR4A1 function rescued FOXG1-dependent death of hippocampal progenitors. Our results reveal differential vulnerability of neuronal subtypes to increased FOXG1 dosage and suggest that activity of a FOXG1/NR4A1 axis contributes to such subtype-specific response.

Newborn granule cells become functionally integrated into the synaptic circuitry of the adult dentate gyrus after a morphological and electrophysiological maturation process. The molecular mechanisms by which immature neurons and the neurites extending from them find their appropriate position and target area remain largely unknown. Here we show that single-cell-specific knockdown of cyclin-dependent kinase 5 (cdk5) activity in newborn cells using a retrovirus-based strategy leads to aberrant growth of dendritic processes, which is associated with an altered migration pattern of newborn cells. Even though spine formation and maturation are reduced in cdk5-deficient cells, aberrant dendrites form ectopic synapses onto hilar neurons. These observations identify cdk5 to be critically involved in the maturation and dendrite extension of newborn neurons in the course of adult neurogenesis. The data presented here also suggest a mechanistic dissociation between accurate dendritic targeting and subsequent synapse formation.

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.

In mammals, adult neural stem cells give rise to new hippocampal dentate granule neurons and interneurons of the olfactory bulb throughout life. The microtubule associated protein Doublecortin (DCX) is expressed by migrating neuroblasts and immature neurons, and is widely used as a stage-specific marker of adult neurogenesis and as a marker to identify neurogenic activity in the adult brain per se. Mutations in the DCX gene have been causally linked to human lissencephalic syndromes. Moreover, embryonic loss of DCX function interferes with neuronal migration and dendritic patterning in a species- and region-specific manner. A putative function of DCX in adult neurogenesis has not been directly explored. Here we show that overexpression of DCX in newly generated dentate granule neurons of the adult mouse brain has no effect on morphological maturation or migration. We also show that micro (mi) RNA-mediated retroviral knockdown of DCX does not alter morphological maturation of adult born dentate granule cells or migration of new neurons in either adult neurogenic niche. Thus, the present data indicate that DCX is dispensable for the development of new neurons in adult mice.

C-terminal Binding Protein 1 (CTBP1) is a ubiquitously expressed transcriptional co-repressor and membrane trafficking regulator. A recurrent de novo c.991C>T mutation in CTBP1 leads to expression of p.R331W CTBP1 and causes hypotonia, ataxia, developmental delay, and tooth enamel defects syndrome (HADDTS), a rare early onset neurodevelopmental disorder. We generated hESCs lines with heterozygote and homozygote c.991C>T in CTBP1 using CRISPR/Cas9 genome editing and validated them for genetic integrity, off-target mutations, and pluripotency. They will be useful for investigation of HADDTS pathophysiology and for screening for potential therapeutics.

Development of oligodendrocytes and myelin formation in the vertebrate central nervous system is under control of several basic helix-loop-helix transcription factors such as Olig2, Ascl1, Hes5 and the Id proteins. The class I basic helix-loop-helix proteins Tcf3, Tcf4 and Tcf12 represent potential heterodimerization partners and functional modulators for all, but have not been investigated in oligodendrocytes so far. Using mouse mutants, organotypic slice and primary cell cultures we here show that Tcf4 is required in a cell-autonomous manner for proper terminal differentiation and myelination in vivo and ex vivo. Partial compensation is provided by the paralogous Tcf3, but not Tcf12. On the mechanistic level Tcf4 was identified as the preferred heterodimerization partner of the central regulator of oligodendrocyte development Olig2. Both genetic studies in the mouse as well as functional studies on enhancer regions of myelin genes confirmed the relevance of this physical interaction for oligodendrocyte differentiation. Considering that alterations in TCF4 are associated with syndromic and non-syndromic forms of intellectual disability, schizophrenia and autism in humans, our findings point to the possibility of an oligodendroglial contribution to these disorders.